DETERMINATION OF THE ANTI BACTERIAL PROPERTY OF SEMI-PURIFIED FLAVONOIDS OF Musa

paradisiaca (Saging na Saba) PEEL AGAINTS Staphylococcus aureus AND Escherichia coli

IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE

FACULTY OF DEPARTMENT OF PHARMACY

SUBMITTED BY:

AYSON, VIARA MAE S.

BAYA, PAMELA V.

CARABBACAN, MARRIANE MYRTELLE MAE D.

RAMOS, FEROZ KYNA B.

FEBRUARY 2020

TABLE OF CONTENTS

ACKNOWLEDGEMENT………………………………………………………………… 6

DEDICATION……………………………………………………………………..………. 7

CHAPTER I

THE PROBLEM AND ITS BACKGROUND

1
INTRODUCTION…………………………………………………….…… 8

Statement of the problem…………………………………………………. 11

Hypothesis………………………………………………………………….. 11

Scope and Limitation…………………………………………………….... 12

Significance of the Study……………………………………………………12

Conceptual Framework…………………………………………………….13

Figure 1: Paradigm of the Study …………………………………………. 15

Definition of terms…………………………………………….…………. . .16

CHAPTER II

REVIEW OF RELATED LITERATURE AND STUDIES

RELATED LITERATURE

Local Literature……………………………………….……………19

Foreign Literature…………………………….…………………….20

RELATED STUDIES

Local Studies…………………………………………………….......23

Foreign Studies……………………………………………………...24

Synopsis……………………………………………………………...26

CHAPTER III

RESEARCH METHODOLOGY

3.1 Research Design……………………………………………………...……. 28

3.2 Research Instruments ……………………………….…………………… 28

3.3 Paradigm of the Data Collection …………………………………………. 30

3.4 Figure 2: Research Procedures ……………………………………………30

3.5 Collection and Preparation of Plant Materials……………………………31

3.6 Figure 3: Test of Saponins………………………………………………… 31

3.7 Figure 4: Test of Flavonoids………………………………………………. 32

2
3.8 Figure 5: Test of Alkaloids…………………………………………………33

3.9 Figure 6: Test for Tannins and Polyphenolic Compound……….……… 33

3.10 Figure 7: Test of Glycosides………………………………………………. 34

3.11 Figure 8: Isolation of the Semi-Purified Flavonoids…………………….. 34

3.6 Research Locale………………………………………………….………… 35

CHAPTER IV

PRESENTATION, ANALYSIS AND INTERPRETATION OF DATA

4.1 Interpretation of Data………………………………………………………36

4.2 Table 1: Organoleptic Testing ……………………………………………..37

4.2 Figure 9: Plant Sample…………………………………………………….. 37

4.3 Figure 10: Collection and Preparation of Plant Sample……………........ 38

4.4 Table 2: Phytochemical screening …….………………………………….. 38

4.6 Table 3: Anti-Microbial Assay of Musa paradisiaca……………………..40

4.7 Table 4: Anti-Microbial Assay of Ciprofloxacin ………………………..... 40

4.8 Figure 11: Anti-biotic disk - Ciprofloxacin (Staphylococcus aureus) ….41

4.9 Figure 12: Anti-biotic disk - Ciprofloxacin (Escherichia coli) ……….... 41

CHAPTER V

SUMMARY OF FINDINGS, CONCLUSIONS AND RECOMMENDATIONS

5.1 Conclusions………………………………………………………………….43

5.2 Recommendation……………………………………………………………44

BIBLIOGRAPHY……………………………………….…………………........................46

APPENDIX

APPENDIX A ………………………………………………………………………49

APPENDIX B.1 ………………………………………………………………….…50

3
APPENDIX B.2 …………………………………………………………………….51

APPENDIX C……………………………………………………………………… 52

APPENDIX D ………………………………………………………………………53

APPENDIX E………………………………………………………………………..54

CURRICULUM VITAE………………………………………….………………………...55

Acknowledgement

The completion of this study will not be a success without the contributing effort of the people who
unselfishly shared their expertise and wisdom to the researchers, without them this study will not be
realized.

4
The preparation of this study was made possible with the exceptional dedication and commitment of
the loving parents of each member of the group.

We are grateful for the invaluable support of Mrs. Floriza De Leon, Ms. Pauline Lyzell G. Francisco,
RPh, MSHSAc, who offered their time in guiding and teaching guidelines and procedures to all the
members of the group.

We wish to thank also the members of the faculty of the College of Pharmacy, Philippine
Rehabilitation Institute, Pampanga Campus who gave us different information and insights which
helped us in creating ideas to come-up with a good outcome of the study. We also want to
acknowledge the opportunity to form a stronger friendship between fellow group mates.

Above all, we would like to give thanks to the God, The Father who always guide and protect us in
every aspect of doing this research. All the glory in His Name.

Dedication

This study is dedicated wholeheartedly to our beloved parents, who were our source of
inspiration and gave us strength when we thought about giving up, providing their moral, spiritual,
emotional and financial support.

To our thesis adviser, teachers, relatives, friends, classmates and group mates who shared
their words of advice and encouragement to complete this study.

And finally, we dedicated this study to the Almighty God, thank you for guidance, energy,
mind power, security, and talents, and for giving us a healthy life. All of these, we offer to you.

5
Ayson, Viara Mae S.

Baya, Pamela V.

Carabbacan, Marriane Myrtelle Mae D.

Ramos, Feroz Kyna B.

CHAPTER 1

THE PROBLEM AND ITS BACKGROUND INTRODUCTION

Musa sp. (Musaceae) also known as banana is a familiar tropical fruit and important source of food in
the world. From its native South western Pacific home, the banana plant spread to India by about 600
BC and later on it spread all over the tropical world. It is possibly the world's oldest cultivated crop.
(PonmuruganKaruppiah and Muhammed Mustaffa, 2013)

All parts of the banana plant have medicinal applications. It helps combat various types of ailments.
Its peel, however, which is not yet widely used in formulation of products, may exhibit antibacterial
and antifungal properties.

Escherichia coli are bacteria found in the environment, foods, and intestines of people and
animals. E. coli is a large and diverse group of bacteria. Although most strains of E. coli are harmless,
others can make you sick. (DOH, USA 2019) Some kinds of E. coli such as E. coli O157:H7, can cause
severe abdominal cramps, vomiting and bloody diarrhea, while others cause urinary tract infections,
respiratory illness and pneumonia and other illnesses. (Charles Patrick Davis, MD, PhD , 2019)

Staphylococcus aureus is a major bacterial human pathogen that causes a wide variety of clinical
manifestations. Infections are common in both community-acquired as well as hospital-acquired
settings and treatment remains challenging to manage due to the emergence of multi-drug resistant
strains such as MRSA (Methicillin-Resistant Staphylococcus aureus).

S. aureus is found in the environment and is also found in normal human flora, located on the skin
and mucous membranes (most often the nasal area) of most healthy individuals. (Tracey A. Taylor;
Chandrashekhar G. Unakal. 2019)

Ciprofloxacin is very active in vitro against most Gram-negative bacteria including Enterobacteriaceae
(especially enteropathogens such as Escherichia coli, Salmonella spp. and Shigella spp.), ciprofloxacin
was used to treat infections caused by Gram negative and Gram positives organisms very successfully
and resistance was very rare. (Oman Med J., 2013)

6
Today, we can list a number of organisms in both hospitals and the community that can prevent
treatment because they are resistant not to just one, but also to numerous kinds of antibiotics. This
study was conducted with high hopes of discovering and developing a new possibility which may help
in the growing health issues of today. Banana as the oldest cultivated plant worldwide makes it easier
to have access with. The peel of the banana is considered waste making it inexpensive.

BACKGROUND OF THE STUDY

Fruit peels have been a valuable source for maintaining human health. The use of fruit peels extracts
for antimicrobial properties can be of great significance in therapeutic treatments. This study aimed
to evaluate the antibacterial activity of banana's peel, extracts of fresh yellow banana peels could be
considered as a good antibacterial agent against both Gram positive and negative bacteria to replace
the synthetic medicines in treatment of diseases caused by these bacteria (Ghani et al., 2013).

Bananas are a highly nutritious fruit, but do they have any medicinal properties beyond their
nutritive value. Among the properties cited:

Flowers: Used to treat dysentery, ulcers, and bronchitis. Cooked, flowers are considered a
good food for diabetics.

Sap: Chemically, banana sap has astringent qualities. In traditional medicine, the sap is used
to treat a wide variety of ailments, including leprosy hysteria, fever, digestive disorders, hemorrhage,
epilepsy, hemorrhoids, and insect bites.

Roots and Seeds: Treat digestive disorders

Peel and Pulp: Scientifically shown to have both antifungal and antibiotic components. These
structures have also been identified as containing the neurotransmitters norepinephrine, serotonin
and dopamine.

E. coli O157:H7 results from its production of Shiga toxin that injures the intestine by sloughing off of
intestinal mucosa cells and results in hemorrhagic diarrhea. The Shiga toxin has systemic effects on
vascular endothelial cells, resulting in vasculitis, and manifests in hemolytic uremic syndrome,
abdominal pain, and rarely, thrombotic thrombocytopenic purpura. E. coli 0157:H7 Shiga toxin
initiates the inflammatory cascade that causes leukocyte aggregation, apoptosis of the affected cells,
platelet aggregation, micro thrombi formation, hemolysis, and renal dysfunction, as the renal
glomeruli have a particular vulnerability to a micro thrombi formation. E. coli 0157:H7 Shiga toxin
effect manifests not only in the kidney, but in its most severe manifestations, it can result in a diffuse
vasculitic injury that affects multiple organ systems and multiple organ failures. (William Gossman;
Abdul Wasey; Philip Salen., 2019)

Staphylococcus aureus are one of the most common bacterial infections in humans and are the
causative agents of multiple human infections, including bacteremia, infective endocarditis, skin and
soft tissue infections (e.g., impetigo, folliculitis, furuncles, carbuncles, cellulitis, scalded skin
syndrome, and others), osteomyelitis, septic arthritis, prosthetic device infections, pulmonary
infections (e.g., pneumonia and empyema), gastroenteritis, meningitis, toxic shock syndrome, and

7
urinary tract infections. Depending on the strains involved and the site of infection, these bacteria can
cause invasive infections and/or toxin-mediated diseases. (Tracey A. Taylor; Chandrashekhar G.
Unakal., 2019)

Due to the increasing cases of antimicrobial resistance, it is viable to search for a new
resource. The purpose of this experimental research is to determine the anti-microbial activity of the
Musa paradisiaca peel and its affectivity against gram-positive and gram-negative bacteria. This study
will provide a new wisdom on the future development of new antimicrobial breed.

Statement of the problem

The main objective of the study is to evaluate the presence of flavonoid present in the
banana peel as antibacterial against Escherichia coli. The study will determine the answer to the
following questions.

1. What is the percentage yield obtained in the semi purified Flavonoids?

2. What are the phytochemical constituents present in the banana peel?

3. What are the organoleptic properties of the plant sample?

4. What are the functional groups present in the plant extract?

5. Is there a significant difference between the positive control (ciprofloxacin) and the
semi-purified flavonoids from the banana peel?

Hypothesis

Null Hypothesis

There is no antibacterial property of Musa paradisiaca peel (Saging na Saba)

extract between Staphylococcus aureus and Escherichia coli.

Alternative Hypothesis

` There is a significant difference between the positive control, Ciprofloxacin

antibiotic disc, from the semi-purified flavonoids isolated from Musa paradisiaca (saging na
saba) peel in terms of zone of inhibition.

8
Scope and Limitation

This research was conducted by the Pharmacy Students of the Philippine Rehabilitation Institute of
Pampanga Campus. This study includes the collection of bananas (Musa paradisiaca) peel followed by
extraction and isolation of the active constituent; flavonoids. Phytochemical screening was
performed by the DOST-Region Philippines. (Regional Gov't. Center Park, Main Road
DiosdadoMacapagal Regional Center, San Fernando, 2000 Pampanga). This study excluded the
biological testing on animals. Moreover, the study is only limited on microbial assay only.

Significance of the Study

Musa paradisiaca is one of the many plant species found in the Philippines. It is an alternative source
of active metabolites or chemicals used in medical treatments. With this discovery, cheaper
medicines made from this plant may be locally available. Thus, it can help alleviate the growing
problems in both the economic and medical sectors.

Agricultural Department.This will help in the development of local plants products in the
Philippines. It will also be of help in producing more of this type of plant that would be great benefits
for our country.

Community Indigents. This study can make the people around the community aware of the
different uses of Musa paradisiaca (Saging Saba). It is not only for mere decoration and consumption
but more importantly can be a cure to common ailments, and used as an anti-bacterial.

Country (Philippines). This will help the government and the country in terms of proper
disposal of trash, to make it beneficial by turning the peels into medicine. This will help and improve
our country by means of agriculture and medicine field.

Department of Health (DOH). This study can contribute in the future programs of the
government especially in the field of medicine. The department of health (DOH) may be aware of the
benefits of using traditional ways and come up with a program that would benefit the public.

Food and Drug Administration. One of the goals of FDA is to provide an effective drug with a
low cost for the public this study of anti-bacterial property of saging na saba can be a big help since it
is very common here in the Philippines and the cost is affordable.

Health Education. This study aims that each student may become more conscious in their
health and personal hygiene. A formulation can be developing such as hand sanitizer.

Medical Technologist. This study will give them guidance and information that they can use
as their reference if the musa paradisiaca extract can cause anti-bacterial effects.

Pharmaceutical Companies. This study could help them classify the type of banana that
should be used. Once they saw the potential there will be an advancement in the investigation and
experiment to evaluate the research that will help in the discovery of a new anti-bacterial drug.

9
 Pharmacy Students. This study is helpful as it will cultivate knowledge and skills on matter of
traditional and alternative medicine. They can use this as a guide in developing alternative methods
of healing using these traditional medicines.

Future Researchers. This study will help the future researchers to be aware and give guidance
and assistance regarding the analysis of anti-bacterial activity present in saging na saba which they
can be use as their future reference.

Conceptual Framework

In this study, the researchers followed the step-by-step procedures stated below: First, is through the
selection of plant sample that contains a constituent with an antibacterial property which is Musa
paradisiaca, next is through the collection of plant sample at Sindalan Market, City of San Fernando,
and Pampanga. The collection is then followed by the authentication of the plant sample at Bureau of
Plant Industry done on the 08th of August 2019. After having certified the plant sample, the
researchers macerated the dried peel of Musa paradisiaca with 80% ethyl alcohol, stand for 48 hours.
This is followed by the extraction of the macerated Musa paradisiacal (Saging na Saba) peel. The
extract was then subjected to different tests – organoleptic, phytochemical, isolation of flavonoids,
and evaluation of the activity of plant sample. In the organoleptic test, the researchers critically
observed the plant’s color, odor, and appearance. In the phytochemical testing, the researchers
submitted their sample at the Department of Science and Technology who confirmed the presence of
its active ingredient through test for flavonoid, test for alkaloids, and test for saponins and then the
procedure for the isolation of flavonoids was done. The evaluation of the antibacterial property of the
semi-purified flavonoids of Musa paradisiaca (Saging na saba) is tested through Microbiological Assay
with the use of Staphylococcus aureus and Escherichia coli.

Figure 1. Paradigm of the Study

10
The paradigm of the study shows the process involved the evaluation of the antibacterial property of
the Semi-Purified flavonoid from Musa paradisiaca peel.

Definition of terms

Agar plate. An agar plate is a Petri dish that contains a growth medium(typicallyagarplus
nutrients) used to culture microorganisms. (Madigan, 2015)

Antibacterial Assay. This is a method for quantitatively determining the concentration of

an antibiotic by its effect in inhibiting the growth of a susceptible microorganism. (Madigan, 2015)

Antibiotic. A chemical substance produced by a microorganism, which has the capacity to

inhibit the growth of or kill other microorganism; antibiotics sufficiently nontoxic to the host are used
in the treatment of infectious disease. (Dorland’s Pocket Medical Dictionary 27th Edition)

Antimicrobial Resistance. Antimicrobial resistance (AMR) is the ability of a microbe to resist

the effects of medication previously used to treat them. This broader term also covers antibiotic
resistance, which applies to bacteria and antibiotics (Madigan, 2015)

Banana. A tropical plant of the genus Musa of the family Musaceae, certain species of which
cultivated for their nutritious fruit. (Webster’s Universal College Dictionary)

Confirmatory Test. are the tests required to confirm the analysis. Confirmatory tests cost
more than simpler presumptive tests so presumptive tests are often done to see if confirmatory tests
are necessary (Alyson Grine; Sarah Rackley, 2016).

Escherichia coli (E. coli): are bacteria found in the environment, foods, and intestines of
people and animals. E. coli are a large and diverse group of bacteria. Although most strains of E. coli
are harmless, others can make you sick. Some kinds of E. coli can cause diarrhea, while others cause

11
urinary tract infections, respiratory illness and pneumonia, and other illnesses. (Satoshi Ishii; Michael
J. Sadowsky, 2017)

Incubator An apparatus for maintaining optimal conditions (temperature, humidity etc.) for
growth and development as one used in the early care of premature infants, or one used for cultures.
(Dorland’s Pocket Medical Dictionary 27th Edition)

Microorganism. A microorganism is a living organism such as bacteria, fungi, and viruses too
small to be seen in the naked eye but visible under a microscope. (Dorland’s Pocket Medical
Dictionary 27th Edition)

Musa Paradisiaca. Is the accepted name for the hybrid between Musa acuminata and Musa
balbisiana. Most cultivated bananas and plantains are triploid cultivars either of this hybrid or of M.
acuminata alone; The name was coined by Carl Linnaeus, the father of modern taxonomy, who
borrowed the name for the genus from Georg Eberhard Rumphius. Linnaeus described the only
banana he was familiar with: a specimen cultivated in George Clifford's glasshouse near Haarlem in
the Netherlands and famous for being the first banana to flower in Europe. In 1736 Linnaeus, named
it Musa Cliffortiana, which is technically a 'pre-Linnean' name. He renamed it Musa paradisiaca, in
reference to the forbidden fruit of paradise. (Häkkinen, et, al 2017)

Phytochemical Screening. This refers to the extraction, screening and identification of the
medicinally active substances found in plants. (https://en.wikipedia.org/)

Residue. A remainder, that remaining after removal of other substances. In biochemistry, that
portion of a monomer that is incorporated in a polymer. (Dorland’s Pocket Medical Dictionary 27th
Edition)

Separatory funnel. A separatory funnel, also known as separation funnel, separating funnel,
or colloquially separatory funnel, is a piece of laboratory glassware used in liquid-liquid extractions to
separate (partition) the components of a mixture into two immiscible solvent phases of different
densities. (Fessenden et, al 2016)

Staphylococcus aureus: is a type of bacteria. It stains Gram positive and is non-moving small
round shaped or non-motile cocci. It is found in grape-like (staphylo-) clusters. This is why it is called
Staphylococcus.; Staphylococcus aureus belongs to the family. Staphylococcaceae. It affects all known
mammalian species, including humans. Further due to its ability to affect a wide range of species, S.
aureus can be readily transmitted from one species to another. This includes transmission between
humans and animals; Minor skin infections, such as pimples, impetigo etc. It may cause boils
(furuncles), cellulitis folliculitis, carbuncles. It is the cause of scalded skin syndrome and abscesses. It
may lead to lung infections or pneumonia. (Ananya MD.2017)

Zone of Inhibition Test. In the Disk Diffusion Antibiotic Sensitivity test (The Kirby-Bauer test) a
thin film of bacteria applied on a plate is subjected to various antibiotics. The Zone of inhibition is a
circular area around the spot of the antibiotic in which the bacteria colonies do not grow. This work
also determines if a particular bacterium is susceptible or resistant to the applied antibiotic using the
calculated Zone of Inhibition and the prescribed standard values. (Bhargav et, al 2016)

12
CHAPTER 2

REVIEW OF RELATED LITERATURE AND STUDIES

This covers the review of literature and studies and synopsis. Almost all species of Banana are
cultivated in the country. It is one of the country’s best agricultural products. All parts of the banana
plant exhibit different medicinal applications. The peel in particular has an anti-fungal and
antibacterial property because of the presence of different secondary metabolites like flavonoids and
alkaloids. In centuries past, history showed that preparations that contains or rich in flavonoids are
used and believed by physicians and lay healers in seek to deal with human diseases. The peel of the
banana is often considered a waste. Discovery of low-end antimicrobial agents is a great need for this
age because of the vast clinical problem; antimicrobial resistance.

LITERATURE

Local

The banana plant is the largest herbaceous flowering plant. The main or upright stem is
actually a pseudo stem, growing from a corm, to a height of 6 to 7.5 meters. Leaves are spirally
arranged, as long as 2.7 meters and 60 cm wide, fragile and easily torn by wind, with the familiar
frond look. Each pseudo stem produces a single stem of bananas; the pseudo stem dies after fruiting,
as offshoots usually develop from the base of the plant. Each pseudo stem produces a single
inflorescence, the banana heart, in a hanging cluster made up of tiers (hands), up to 20 fruit to a tier.

Cardaba bananas are botanically known as part of Musa acuminate x balbisiana (ABB Group) ‘Saba’
and a member of the Musaceae family. The Cardaba banana is also known as Cardava and
sometimes mistakenly called saba. The Cardaba banana is native to the Philippines and along with
the saba are the most common cooking banana cultivars grown there today.

Saba' is the name given in the Philippines to a cooking banana belonging to the Saba subgroup. Its
classification at the genome group level has been disputed ever since it was suggested to be a triploid
derived from crosses between Musa balbisiana only. Whereas some molecular analyses seem to
support a BBB genome group, morphologicaltraits characteristic of Musa acuminata, such as curly
bracts, place it among the ABB. On the other hand, if backcrossing occurred, it is possible that
backcrosses with M. balbisiana might have led to the substitution of M. acuminata genes by M.
balbisiana ones, leaving only remnants of the original M. acuminata genome. 'Saba' is especially
popular in the Philippines.Cruz, F. S. (2015). Farmers' Handbook on Introduced and Local Banana
Cultivars in the Philippines.

Foreign

13
 Banana (Musa paradisiaca, family Musaceae) is a central fruit crop of the tropical and
subtropical regions of the world. It is possibly the world’s oldest cultivated crop. It is found that Musa
sapientum consists of glycosides, alkaloids, tannins, and flavonoids.

Flavonoid is of the constituent that exhibit antibacterial activity. It is the largest group of
phytonutrients. Flavonoids are part of the polyphenol class of phytonutrients. Polyphenols have
historically been used in Chinese and Ayurvedic medicine and are associated with skin protection,
brain function, blood sugar and blood pressure regulation, in addition to antioxidant and anti-
inflammatory activity (Kumar et al., Global Healing Center, 2012)

Musa paradisiaca L. (Family: Musaceae), commonly known as plantain is a perennial tree-like herb
widely distributed in the tropics. M. paradisiaca is still largely unexplored source for the development
of new drugs. Globally, about 85% of all medications for different diseases are derived from plants.
Hence, the aim of the present review on M. paradisiaca is to collect all the work done on this
important plant in one place, to facilitate the re- searchers to explore further on this plant.
(Lakshmi, Vijai 2016)

It is a tall, robust herb, and the plant portion above the ground is a false stem (pseudo-stem)
consisting of concentrically formed leaves, from the center of which develops the inflorescence stalk.
The rhizome or true stems underground. Near the tip of the flower stalk are several groups of sterile
male flowers subtended by brilliant purple bracts.(Lakshmi, Vijai 2017)

The lower female flower clusters on the same stalk and give rise to the fruit. The whole plant as well
as specific parts (Flowers, banana bracts, ripe, unripe fruits, leaves and stems) of plant extract and its
active constituents have been used for the treatment of large number of human ailments.
Traditionally the plant has been used for different purposes such as abscess, alopecia, burns, cancer,
cataplasm, diabetes, diarrhea, dog bites, snake bite, dysentery, dyspepsia, fracture, gangrene,
hematuria, hemiplegia, hemoptysis, hemorrhage, hypertension, lizard bites, marasmus, migraine,
ring- worm, shingles, smallpox, syphilis, tuberculosis, tumor, uremia, otalgia, psoriasis,
urticaria, warts and wounds. Similarly, the pulp has antiulcer, wound healing, hair growth
promoting, analgesic, and antioxidant property and hepatoprotective activities.

E. coli O157:H7 is transmitted to humans primarily through consumption of contaminated foods, such
as raw or undercooked ground meat products and raw milk. Faecal contamination of water and other
foods, as well as cross-contamination during food preparation. An increasing number of outbreaks
are associated with the consumption of fruits and vegetables (including sprouts, spinach, lettuce,
coleslaw, and salad) whereby contamination may be due to contact with faeces from domestic or wild
animals at some stage during cultivation or handling. STEC has also been isolated from bodies
of water (such as ponds and streams), wells and water troughs, and has been found to survive for
months in manure and water-trough sediments. Waterborne transmission has been reported, both
from contaminated drinking-water and from recreational waters. (William Gossman; Abdul Wasey;
Philip Salen., 2019)

Staphylococcus aureus is a human commensal that can also cause systemic infections. This transition
requires evasion of the immune response and the ability to exploit different niches within the
host. However, the disease mechanisms and the dominant immune mediators against infection are
poorly understood. Previously it has been shown that the infecting S. aureus population goes through

14
a population bottleneck, from which very few bacteria escape to establish the abscesses that are
characteristic of many infections. (Pollitt et al., 2018)

Despite this medicinal importance of M. paradisiaca, it is usually cultivated for food. It is consumed as
an energy yielding food and desert. Fruits are an important contribution to the diets of many people.
India is the largest producer of banana. The states of Maharashtra and Gujarat in Western India,
Karnataka in Southern India and Assam in the northeast are large banana growers. (Lakshmi, Vijai
2015)

STUDIES

Local

The ethanol extract of peel of Musa paradisiaca and its aqueous fraction were investigated for
antibacterial potentials against Gram-positive and Gram-negative organisms of clinical
importance. The extract showed activity by inhibiting the growth of these organisms and causing a
reduction in their viable count with time while showing only a moderate toxicity in mice.
These findings provide scientific explanation for the use of this plant leaf in traditional medicines and
shows that it can be exploited as a possible source of new antibiotics. (Asuqou, 2016)

The agar well diffusion method was used to carry out this test. 0.1 ml of 1:1000 dilution of test
organism for Gram-positive and 1:5000 dilution of test organism for Gram-negative were introduced
into labeled sterile Petri dishes. Twenty milliliters of the cooled nutrient agar medium were
aseptically poured into each Petri dish and gently swirled to mix. The plates were allowed to set and
wells were created using sterile 4-mm cork borer. Different concentrations (200–12.5 mg/ml) of the
extract diluted with water was introduced into the different wells and labelled appropriately. The
Petri dishes were allowed to stand for 20 min before incubation at 37°C for 24 h. The diameters of the
zones of growth inhibition were measured in millimeters (mm).

The aqueous fraction of the ethanol extract exhibited higher antimicrobial activities compared
to the crude ethanol. This is because the secondary metabolites being polar compounds like water
have been pooled into the aqueous fraction during fractionation. S. Aureus had a wider zone of
inhibition than others, indicating a possibility in the use of the plant to treat infections due to the
organism which are on the increase. The antimicrobial properties of plant

It indicates that the crude and the aqueous fraction of the ethanol extract of Musa Paradisiaca peel
possess antibacterial activity. The aqueous fraction showed greater activity than the crude extract
against the test organisms. The concentration of Musa paradisiaca peel extract required to arrest the
growth of test organisms is relatively less. The time–kill curves clearly show the ability of the extract
to reduce microbial population exponentially with time. As such, the antibacterial activity of M.
Paradisiacal can be exploited for the treatment of infections caused by the test organism at
relatively low doses to avoid toxicity. (Asuqou, 2016)

Foreign

Banana is one of the most popular fruits distributed all over the world. Traditional, the fruit, Stem
juice, flowers of banana plants wereused for treating diarrhea(unripe), dysentery, menorrhagia,
diabetes, antilithic, antiulcerogenic, hypoglycemic,hypolipidemic,anti-oxidantactions

15
andiinflammation, pain & snakebite.In order to justify the ethno medicinal claims of the ethanol
extracts of Musa paradisiaca.

Minimum Inhibitory Concentration was determined for extracts that showed ?7 mm diameter growth
inhibition zone. The test was performed using agar well diffusion methods. In agar well diffusion, the
extracts (500 µg/mL) was serially diluted as 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, 1:128 and 1:256 to bring
250 µg/mL, 125 µg/mL,62.5 µg/mL, 31.25 µg/mL, 15.63 µg/mL, 7.81 µg/mL, 3.95 µg/mL and
1.95µg/mL concentrations, respectively. The extract was then aseptically introduced into an agar well.
The inhibition zone was measured after 24 h incubation at 37 °C and the minimum concentration that
inhibited growth was considered as MIC value of the extracts. (Karuppiah, Mustaffa, 2013)

Minimum Bactericidal Concentration was determined by sub-culturing the samples having a value of
lesser or equal to MIC value. The highest dilution (lesser concentration) that yielded no single
bacterial colony was taken as MBC. (Karuppiah, Mustaffa, 2013)

The evaluation of antibacterial activity of non-polar (hexane) and polar (ethyl acetate, methanol)
extracts of the leaves of Musa sp. against seven gram-negative and two gram-positive bacteria were
studied using agar well diffusion method. The results revealed variability in the inhibitory
concentration of each extracts against a panel of pathogenic bacteria. (Karuppiah, Mustaffa,
2013)

The diameters of growth inhibition zone were in the range maximum of 18.6±0.5 mm and minimum
of 8.0±0.5 mm whereas the standard drug, chloramphenicol had higher zones of inhibition ranging
from 11.1±0.3 mm to 22.4±0.20 mm. (Karuppiah, Mustaffa, 2013)

With its broad spectrum of antibacterial activity and ability to achieve therapeutic concentrations in
most body fluids and tissues, Ciprofloxacin has proved useful in the treatment of a wide variety of
infections. (Oman Med J. , 2013)

Clinical and bacteriological cure rates for uncomplicated urinary tract infections were >90% with 3- to
7-day Ciprofloxacin regimens (500 mg/day; >90% of patients were female), and were slightly lower
with single-dose regimens. A3-day course of low-dose Ciprofloxacin (100mg twice daily) was as
effective as cotrimoxazole(trimethoprim-sulfamethoxazole) [3- and 7-day regimens], nitrofurantoin (7
days) and Ofloxacin (3 days) in this indication. Cure rates ranged from 76 to 96% in complicated
urinary tract infections, and Ciprofloxacin was as effective as cotrimoxazole, ceftazidime,
aminoglycosides and other fluoroquinolones. (Oman Med J. , 2013)

Grand naine and Musa paradisiaca. Potable were screened for potential antibacterial activity using
agar well diffusion method against 8 clinical st rains.Extracts of M. paradisiaca showed a
significant level of bacterial inhibition against Proteus vulgaris (19.78±0.40), Pseudomonas
aeruginosa (19.67±0.41) and Staphylococcusaureus (17.44±0.50), moderate activity against
Salmonella typhi (16.89±0.48), Salmonella paratyphi (16.67±0.37), Klebsiellapneumoniae
(15.56±0.50),Bacillus subtilis (15.11±0.42) and very less against Escherichia coli (09.11±0.39).

Synopsis

16
 The research study is formed for the purpose of using the disposable part of a plant as a
remedy/ medicine, that will help the community and also providing an additional knowledge to the
indigenous plant. In order to achieve that goal, the researchers will perform an experimental research
that will be provided with quantitative and qualitative variables/ factors.

As for that, the researchers conduct in-vitro tests which consist of phytochemical
screening in order to detect the pharmacological benefits of the plant sample (Musa paradisiaca), and
anti-microbial tests that will identify the effectivity of our plant extract against two microorganism
(Staphylococcus Aureus and Escherichia coli) that holds a large broad spectrum on the category of
gram positive and gram negative. Positive control (Ciprofloxacin) was chosen consideringits
spectrum, effectivity and action against the two said bacteria.

According to the studies stated, the diameters of growth inhibition zone were in the range maximum
of 18.6±0.5 mm and minimum of 8.0±0.5 mm standard using the drug chloramphenicol whereas in
this study, we conducted the testusing ciprofloxacin that is alsovery active in in vitro against most
Gram-negative bacteria which gives us the positive result as well.

CHAPTER 3

RESEARCH METHODOLOGY

This chapter addresses the research methods and procedure provides a detailed overview of how the
study was conducted and details the specific steps and tasks subject’s, the analysis of data and the
numerical treatment

Research Design

The researchers study utilized the experimental research design to test the Musa paradisiaca (Saging
na Saba) peel anti-bacterial properties. Experimental research is any science-based study in which a
set of variables is kept constant while the other set of variables are calculated as the object of the
experiment. (Bhat A., 2019)

17
Research Instruments.

The following instruments were used to conduct the research:

Agar plate. A plate is used to culture microorganisms (Escherichia coli.)

Antibiotic disk. This antibiotic disk is used for bacteria sensitivity testing and allow you to test
up to two different bacteria cultures with four different antibiotics.

It served as the positive control.

Ciprofloxacin antibiotic disc. Served as the positive control for the antibacterial assay.

Dehydrator. A device that is used to dry, or dehydrate. Utilizing a built-in fan and low amounts
of heat.

Escherichia coli. Also known as E. coli, is a gram, negative, rod-shaped, coliform bacterium
that is commonly found in the lower intestine of warm-blooded organisms. It is one of the most
frequent causes of many common infections, including urinary tract infection (UTI), traveler’s
diarrhea, bacteremia, etc.

Incubator. It is a device used to grow and maintain microbiological Cultures or cell cultures.
The incubator maintains optimal temperature

Microbes. Staphylococcus aureus and Escherichia coli was the microorganism subjected to
microbiological testing.

Osteorizer. It is used to grind the plant sample into fine powder.

Phytochemical Screening. The phytochemical screening was conducted in order to identify

the presence of possible constituent’s presents in the peel of Musa paradisiaca such as alkaloids,
saponins, glycoside, steroids, and flavonoids. Color reactions were observed with subject to different
test reagents.

Percentage Yield

Separatory funnel. An apparatus used in liquid-liquid extractions to separate (partition) the

components of a mixture into two immiscible solvent phases of different densities.

Staphylococcus aureus. Gram-positive round shape bacterium that is a member of the

firmicutes, and usually found in upper respiratory tract and in the skin. It can cause a range of
illnesses, from minor skin infections such as pimples, impetigo, boils, abscesses etc. and life-
threatening diseases such as pneumonia, meningitis, sepsis etc.

18
Data Collection and Procedures

Figure 2: Research Procedures

Determination of the antibacterial property of the Musa paradisiaca (saging na saba) peel.

Collection and Preparation of Plant Materials

Collection of Plant Material

The plant sample, which consists of the peel of Musa paradisiaca, will be collected
from Sindalan Market, City of San Fernando, Pampanga. The peel of Musa paradisiaca is

19
washed, cleaned and dried using a dehydrator for 18 hours. To reduce the size of the cleaned peel
of Musa paradisiaca, it will be osterized until smaller particles is acquired.

Plant extracts

Ethanolic extract

In order to obtain the crude extracts, around 100g of peel of Musa paradisiaca was
macerated in enough 80% ethanol for 48 hours. The sample was filtered using filter paper and the
residue was discarded. And kept in an air tight glass container and reserved for the preliminary
test and for the isolation of semi purified flavonoids.

Phytochemical Screening

The phytochemical screening was conducted in order to identify the presence of possible
constituent’s presents in the peel of Musa paradisiaca such as alkaloids, saponins, glycoside,
steroids, and flavonoids. Color reactions were observed with subject to different test reagents.

Figure 3: Test for Saponins

Liebermann-Burchard Test

The 10 mL of the crude extract was evaporated to dryness using a water bath. Cool to room
temperature. 10 mL of hexane was added to the cooled residue, stir for few minutes. Allow the
solution to settle and decant the supernatant liquid. The procedure was repeated until the hexane
has removed most of the color of the extract. Discard hexane extract. 10 mL of chloroform was added
to the residue for 5 minutes, decant into a test tube containing about 100 mg of anhydrous sodium
sulfate. Shake and passed through a filter paper. The filtrate was divided into two clean and dry test
tubes, use one portion as the standard. To one portion, add 3 drops of acetic anhydride and mix
gently. Then, add 1gtt of conc. Sulfuric acid and mix gently. Observed any change in color over a
period of 1 hour and compare with the standard. Sterols and triterpenes give a range of colors from
blue to green, red, pink, purple or violet.

Figure 4: Test for Flavonoids

Bate-Smith and Metcalf Test

The 10mL of the crude extract was evaporated to incipient dryness over a water bath, cool to room
temperature. The residue was triturated with 10mL petroleum ether and decant. Titration of the
residue was with fresh volumes until the solvent is almost colorless. Discard the petroleum ether.
Dissolved the defatted residue in 20 mL of the 80% ethyl alcohol and filter off the insoluble residue.
Divide the extract into three portions, for Bate-Smith and Metcalf, for Willstatter “Cyanidin” Test and
for standard.

20
 Test for Leucoanthocyanins: Bate-Smith and Metcalf Method

The 0.5mL of conc. HCl was added and observed for any changed in color. Warm the
test tube on a water bath for 15 minutes. Observed any change in color within an hour and compare
with the standard. A strong red or violet color indicates the presence of leucoanthocyanins.

Test for ?-benzopyrone nucleus: Willstatter “Cyanidin” Test

The 0.5mL of concentrated HCl and 3-4 pieces of magnesium turnings was added to
the extract. Observed any changed in color within 10 minutes and compared with the standard. It
was diluted with equal volume of water and added 1mL of octyl alcohol. Shaken and allowed
standing. The colors ranging from orange to red to crimson and magenta, and occasionally to
green to blue may be observed.

Figure 5: Test for Alkaloids

Mayer’s Test

The 2mL of the crude extract was evaporated to a syrupy consistency on an evaporating dish over a
steam bath. The 5mL of 2M HCl to the conc. was added to the extract and stir while heating for about
5 minutes. Adding 5mL of 2M HCl again to wash the extract filter and collect the filtrate to the final
volume of 5mL.1mL of the filtrate in a separate test tube and add few gtts of Dragendorff’s reagent.
With Mayer’s reagent, a positive test is indicated is indicated by a white precipitate and with
Dragendorff’s reagent; a positive test is indicated by an orange precipitate.

Figure 6: Test for Tannins and Polyphenolic Compounds

Ferric Chloride Test

The 10mL of crude extract sample was evaporated to incipient dryness over a water bath and cooled.
Then, the residue was extracted with 20mL of hot water and cooled. Adding 5gtts of 10% NaCl
solution to salt-out undesirable constituents and then filtered.Then 3gtts of ferric chloridesolution
was added. A blue-black color indicates the presence of hydrolysable tannins while a brownish-green
color may indicate the presence of condensed tannins.

Figure 7:Test for Glycosides

Keller- Killani Test

Treat 5ml of each extract with 2 ml of glacial acetic acid containing one drop of ferric chloride
solution. Underlay the mixture with 1 ml of conc. sulfuric acid. A brown ring of the interface indicates
a deoxy sugar characteristic of cardenolides. A violet ring mayappear below the brown ring, while in
the acetic acid layer, a greenish ring may form just gradually throughout thin layer.

21
Figure 8: Isolation of the Semi-Purified Flavonoids

Research Locale

The peel of Musa paradisiacapeel (Sagingna Saba) was bought from Sindalan Market, City of San
Fernando Pampanga and was processed at Philippine Rehabilitation Institute Pharmacy Laboratory,
Pampanga Campus serve as a plant sample. The laboratory tests and the Microbiological Test were
performed at DOST-Region Philippines. (Regional Gov't. Center Park, Main Road DiosdadoMacapagal
Regional Center, San Fernando, 2000 Pampanga)

CHAPTER 4

PRESENTATION, ANALYSIS AND INTERPRETATION OF DATA

This chapter covers the results and outcomes including the results for phytochemical
screening, Confirmatory test for flavonoids, Microbiological assay, organoleptic test and solubility
test.

The study focused on the antibacterial activity of the semi-purified flavonoids from the peel
of Musa paradisiaca (Saging na saba).

Interpretation of Data

Percentage Yield

The collected semi-purified flavonoids were weighed and the percentage yield is computed
based on the formula:

22
 The percentage yield obtained in the semi purified flavonoids is 55.36%. This shows that it
yields enough percentage to have a positive result.

Organoleptic Testing

Table 1. Results of the Organoleptic testing of the Musa paradisiacal extract.

TESTRESULTColorBrownishOdorSweet Coffee-like odorAppearanceClear brown solution

As seen above, the results showed the organoleptic tests employed on the plant sample. This
includes the odor, color and appearance of the Musa paradisiaca. It was observed that the extract has
a dark brown color and a coffee-like odor.

Figure 9: Plant sample

Figure 10: Collection and Preparation of Plant Sample

The plant samples were gathered at Sindalan Market, City of San Fernando, Pampanga. The
researchers dried the peel of Musa paradisiaca peel (Sagingna Saba) using a dehydrator for 18 hours
at 30°C.

23
Active Constituent Identified in the Phytochemical Screening

Table 2: Phytochemical screening of the constituents of the Plant Sample.

Test ParameterMethod ResultSignificanceAlkaloidsMayer / Meyer Test

Detected

Alkaloidal compounds were presentAlkaloids -- (antimalarial, antiasthma, anticancer, cholinomimetic,

vasodilator, antiarrhythmic, analgesic, antibacterial,anti-hyperglycemic activities, psychotropic and
stimulant)

DETECTED- had a wide range variety of pharmacologic effect but the antibacterial property is the
most significant on the plant sampleAnthraquinonesBornträger Test

Not Detected

Anthraquinone compounds were not presentAnthraquinones- (Stimulant cathartics)

NOT DETECTED- will not affect the tone of smooth muscle in the wall of large intestines. Cardenolides

and BufadienolidesKeller-Killani Test

Not Detected

Glycoside compounds were not presentCardenolides — (branch of cardiac glycosides that treats
cardiac related problems)

NOT DETECTED — this will not affect the heart since it is not present on the sample.

Bufadienolides — (widely used as treatment in cardiac dysfunction)

NOT DETECTED — it will not affect the heart since the compound isn’t presentFlavonoidsBate-Smith
and Metcalf Test

Detected

Anthocyanins were present

Flavonoids (anti-inflammatory, anti-mutagenic and anti-carcinogenic properties)

DETECTED-can contain properties that can avoid inflammation, mutagenesis and can fight
carcinogenic properties.Tannins

And Polyphenolic CompoundsFerric chloride Test

Not Detected

Hydrolysable tannins were not present

24
Not Detected

Condensed tannins were not presentTannins- (astringent, antiseptic)

NOT DETECTED- will not affect the GIT and skin pores.

Polyphenolic compounds (Strong antioxidant property)

NOT DETECTED- cannot prevent or slow damage to cells caused by free radicals.SaponinsFroth Test

Detected

Saponins were presentSaponins — used for detergent and hemolytic property, antimicrobial
properties, improve immune function

DETECTED— can be used in the production of soaps, guarding your body against fungi, bacteria and
viruses, stimulates the production of T-cells.In the Phytochemical screening of the constituents of the
Plant Sample shows positive results of Alkaloid, Flavonoids, and Saponins.

Table 3: Anti-Microbial Assay of Musa paradisiaca (saging na saba) peel extract.

Test ParameterMethodStandard Guide LevelResultAnti-Microbial MethodTM-408

Anti-Microbial Assay Procedure

(Filter Disk Method)

Guevarra 2005 and Benson: Microbiological Applications Laboratory Manual/ 8th Edition 2001

None Available

SUSCEPTIBLE for Gram (+) positive and Gram (-) negative bacteriaThe antibacterial property of the
semi-purified flavonoids from Musa paradisiacapeel were evaluated by the disc diffusion method
against Escherichia coli and Staphylococcus aureusfound to be inactive in both gram positive and
gram-negative bacterial strains.

Test ParameterMethodStandard Guide LevelResultAnti-Microbial MethodTM-408

Anti-Microbial Assay Procedure

(Filter Disk Method)

Guevarra 2005 and Benson: Microbiological Applications Laboratory Manual/ 8th Edition 2001

None Available

25
RESISTANT for Gram (+) positive and Gram (-) negative bacteriaTable 4: Anti-Microbial Assay of
Ciprofloxacin

The Anti-Microbial assay of Ciprofloxacin shows VERY ACTIVE for Gram (+) positive and Gram (-)
negative bacteria.

Figure 11: Anti-biotic disk - Ciprofloxacin (Staphylococcus aureus)

The picture above shows the resistance in the inhibition

of Ciprofloxacin against Staphylococcus aureus.

Figure 12: Anti-biotic disk – Ciprofloxacin (Escherichia coli)

The picture above shows the resistance in the inhibition

of Ciprofloxacin against Escherichia coli.

CHAPTER V

SUMMARY OF FINDINGS, CONCLUSIONS AND RECOMMENDATIONS

26
The research on the semi-purified flavonoids from Musa paradisiaca peel extract (Saging na Saba)
was to determine if the plant has potential to antibacterial activity. Due to the fact that the literature
on the Musa paradisiaca peel (Saging na saba) peel is vast, and only a few studies are focused on
Musa paradisiaca (Saging na saba), this research was chosen and conducted to gain information and
to prove or determine its antibacterial property.

The following findings were based on the results obtained.

1. For the Organoleptic testing, the Musa paradisiaca peel extract, was described to physical testing.

The Musa paradisiaca peel extract has a brownish color.

It also has a sweet coffee-like odor

The extract also looks like a clear brown solution.

2. The analysis on phytochemical screening of Musa paradisiaca peel (Saging na Saba) extract is to
identify different constituent present in the extract, such as:

i. Flavonoids

ii. Alkaloids

iii. Saponins

3. The Department of Science and Technology (DOST), Region 3 conducted the test for the Musa
paradisiaca peel (Sagingna Saba) extract an antibacterial property against Staphylococcus aureus and
Escherichia coli. The results show a <10mm zone of inhibition indicating a negative anti-bacterial
property. Therefore, the result of the test is negative.

4. The implication of this study in the field of pharmacy is for the hope to find a variety of an effective
anti-bacterial property that would be cost efficient and effective. This study will further help the
future researchers and other tropical nation in terms of research. Moreover, this will give them
insight and knowledge in order to develop a new anti-bacterial drug.

Conclusion

1. Based on the performed test and experiments using the extract through phytochemical screening,
Antimicrobial assay the researchers concluded that, the Musa Paradisiaca peel contained different
active constituents Saponins, and Flavonoids, Alkaloids through phytochemical screening.

2. The organoleptic test that was performed gives thesaging na saba extract a brownish color, a sweet
coffee-like odor and looks like a clear brown solution. Hence, having a darker brown to black solution
yields to have a possible negative result for the phytochemical testing.

27
3. In the analysis of the phytochemical screening of saging na saba. The constituents Alkaloids,
Saponins and Flavonoids were detected. This shows that the extract contains an astringent, detergent,
hemolytic, anti-oxidative and anti-inflammatory property.

4.The result of antimicrobial assay conducted revealed that Musa Paradisiaca (Banana peel) has zone
of inhibition with Gram positive and Gram-negative bacteria thus, making the result of antimicrobial
assay conducted revealed that the Musa paradisiaca, (sagingnasaba) does not exhibit antibacterial
property against Staphylococcus aureus and Escherichia coli, opposing the international study.

Recommendations

The following are recommended for the future researchers:

1. The use of other variety of species of the plant. In addition to this, the researchers would
like to recommend the formulation of new studies regarding the anti-bacterial property of the
plant.

2. Discoloration of the extract is preventable and manageable by the proper handling

and preparation of the sample.

3. Try to either use a lower or higher amount of different solvent that would be more
compatible to yield a purer extract from the plant.

4. Increased the number of experimental trials that will give positive results through the
use of in vitro process.

5. Try testing the saging na saba extract to different species of gram negative and positive
microorganisms.

6. To further study about the other therapeutic use of the plant parts such as leaves,
roots, seeds, pulp and flowers.

7. To try conducting other tests and using laboratory equipment’s accurately and review
its efficacy properly.

8. To try adjustments in maceration such as using different volumes or percentage of the

solvent.

9. The researchers recommend to research for an effective formulation to obtain the

anti-bacterial property of Musa paradisiaca and to further increase its potency.

10. Conducting an in-vivo tests on the laboratory using guinea pigs or mice for further
results.

11. Quality control in doing the whole experiment should be strictly utilized to assure
positive result.

12. Use an autoclave for the maceration of the plant sample.

28
BIBLIOGRAPHY

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PE 1628. W49 (2001) Webster’s Universal College Dictionary, Ref 2237

PE 1591 M55 (1991) The Merriam-Webster Thesaurus, Ref 1483

QR 46 J32 (2010) Jawetz, Melnick & Adelberg’s Medical Microbiology 25th Edition, Med 2064

R 121 D67 (2004) Dorland’s Pocket Medical Dictionary 27th International Edition, Ref 066

Farmer’s Handbook on Introduced and Local Banana Cultivars in the Philippines

JOURNALS AND ARTICLES

Agarwal, P K. Evaluation of wound healing activity of extracts of plantain banana (Musa

sapientumvar.paradisiaca) in rats.

Asuquo EG, Udobi CE. Antibacterial and toxicity studies of the ethanol extract of Musa paradisiacal.
Cogent Biology.2016; 2(1). oi:10.1080/23312025.2016.1219248.Cowan, M. Plant Products as
antimicrobial Agents. Retrieved October 28, 2016

Barry L. Arthur of Measuring zone of inhibition with the Bauer-Kirby Disk susceptibility Test,
University of California-Davis, Medical Canter, Sacramento, California 95817; University of
Washington, Harborview Medical Center, and Journal of Clinical Microbiology. Vol. 10, No. 6

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Cruz, F. S. (2015). Farmers' Handbook on Introduced and Local Banana Cultivars in the Philippines.

Egbuono, A. (2016, August 11). Comparative Evaluation of the Proximate Composition and
Antibacterial Activity of Ground Musa paradisiaca (Plantain) Peels and Leaves.

Ehiowemwenguan, G. , www.iosrphr.org Volume 4, Issue 8 (August 2014), PP. 18-25Antibacterial and

phytochemical analysis of Banana fruit peel., Department of Microbiology, University of Benin, P.M.B
1154. Benin City, Nigeria.

Kapadia, SurajPremal MD. Contemporary Clinical Dentistry Detection of Antimicrobial activity of

Banana peel (Musa paradisiacal L.) on Porphyromonasgingivalis and
Aggregatibacteractinomycetemcomitans: An in vitro assay;

Toting, John Paul Tolentino. 2016, Formulation of Hand Sanitizer Gel using the Semi-Purified
Flavonoids from the outer coverings of the Red Creole variety of Allium cepa Linn.: A preliminary
investigation, Department of Pharmacy, Centro Escolar University- Malolos Km. 44 Mc Arthur
Highway, City of Malolos, Bulacan, JAASP

T.P Tim Cushnie, Andrew J. Lamb, International Journal of Antimicrobial Agents (2005) 343-356, Anti-
microbial activity of Flavonoids, School of Pharmacy, The Robert Gordon University, School hill,
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Comparative Evaluation of Phytochemical and Antimicrobial (n.d.) Retrieved July 28, 2016.
Mutagens.co.in/jgb/vol.02/1/03.pdf

HappiEmaga T, Bindelle J, Agneesens R, Buldgen A, Wathelet B, Paquot M. Ripening influences

banana and plant

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Lakshmi, Vijai. NATURAL PRODUCTS: An Overview of Musa paradisiaca Linn. Vol. 11 Issue 4, ISSN:
0974-7508, 2015 [105-109] Cited:

Resistance Pattern of Ciprofloxacin Against Different Pathogens Oman Med J. 2015 Oct; 25(4): 294–
298.doi: 10.5001/omj.2010.85

Appendix A

Certification of Musa paradisiaca from BPI

Appendix B.1

Certificate of Analysis: Phytochemical Screening

Appendix B.2

Appendix C

Certificate of Analysis: Anti-Microbial Assay of Musa paradisiaca

31
Appendix D

Certificate of Analysis: Anti-Microbial Assay of Ciprofloxacin

Appendix E

Certificate of English Editing

AYSON, VIARA MAE S.

#268 Sinura Ave. Sinura, Porac, Pampanga

32
salunga.viara22@gmail.com

Personal Background

Date of Birth: October 21, 1998

Age: 21 years old

Place of Birth: Angeles City, Pampanga

Nationality: Filipino

Civil Status: Single

Name of Father: RamilAyson

Occupation: Company Driver

Name of Mother: Olivia Ayson

Occupation: Housewife

Educational Background

Tertiary: Philippine Rehabilitation Institute Inc.

Bachelor of Science in Pharmacy

Secondary: Holy Trinity Manibaug, Paralaya

Primary: Sinura Elementary School

BAYA, PAMELA V.

Empresa Homes Calulut, Pampanga

bayapamela1013@gmail.com

33
Personal Background

Date of Birth: September 13, 1998

Age: 21 years old

Place of Birth: Floridablanca, Pampanga

Nationality: Filipino

Civil Status: Single

Name of Father: Alan Baya

Occupation: Crane Operator

Name of Mother: Baby Pacifica Baya

Occupation: Housewife

Educational Background

Tertiary: Philippine Rehabilitation Institute Inc.

Bachelor of Science in Pharmacy

Secondary: Pampanga High School

Primary: Magliman Elementary School

CARABBACAN, MARRIANE MYRTELLE MAE D.

#175 Gaffud St. TaggappanEchague, Isabela

marrianemyrtellemae@gmail.com

34
Personal Background

Date of Birth: June 10, 1996

Age: 23 years old

Place of Birth: Santiago City, Isabela

Nationality: Filipino

Civil Status: Single

Name of Father: Robert Carabbacan

Occupation: Deceased

Name of Mother: Victoria Carabbacan

Occupation: Business Woman

Educational Background

Tertiary: Philippine Rehabilitation Institute Inc.

Bachelor of Science in Pharmacy

Secondary: University of La Salette

Primary: Echague West Central School

RAMOS, FEROZ KYNA B.

MauliSta.BarbaraBacolor, Pampanga

ramosferoz08@gmail.com

35
Personal Background

Date of Birth: April 08, 1996

Age: 23 years old

Place of Birth: Guagua Provincial Hospital

Nationality: Filipino

Civil Status: Single

Name of Father: Ferdinand Ramos

Occupation: Civil Engineer

Name of Mother: Paulita Ramos

Occupation: Housewife

Educational Background

Tertiary: Philippine Rehabilitation Institute Inc.

Bachelor of Science in Pharmacy

Secondary: Pampanga High School

Primary: San Fernando Elementary School

36