Antibacterial activity of Turmeric Leaf (Curcuma longa Linn.

) Ethanolic

Extract against Staphylococcus aureus

___________________________

A Research Paper

Presented to

The Faculty of Liceo De La Salle Senior High School

______________________________

In Partial Fulfillment
Of the Requirements for
Capstone Project

______________________________

Aposaga, Keanna Yasmin P.

Cruz, Aliya Margaret M.
Gamab, Ma. Cristina R.
Hilado, Lance Kristoffer T.
Libo-on, Janine Alexa P.
Pacheco, Roberto Matteo N.

STEM 12-F
______________________________

April 2024
 APPROVAL SHEET

The research entitled "ANTIBACTERIAL ACTIVITY OF TURMERIC LEAF(Curcuma

longa Linn.) ETHANOLIC EXTRACT AGAINST Staphylococcus aureus" presented by
KEANNA YASMIN P. APOSAGA, ALIYA MARGARET M. CRUZ, MA. CRISTINA R.
GAMAB, LANCE KRISTOFFER T. HILADO, JANINE ALEXA B. LIBO-ON, AND
ROBERTO MATTEO N. PACHECO in partial fulfillment of the requirements for Capstone
Project of Liceo De la Salle-Senior High School of the University of St. La Salle has been
evaluated and approved by the panel evaluators.

PANEL OF EVALUATORS

KENNETH S. PARRA, RN, LPT. DHARCEE L. PELAGIO, LPT

Panelist Panelist

MARIA ANGELA A. LEONOR, LPT, Ph.D.

Adviser

ACKNOWLEDGEMENT
 The researchers would like to express their gratitude for the guidance of the
following individuals who assisted with the development in accomplishing the research:

To the research adviser, Dr. Maria Angela Leonor for her share of knowledge
regarding their study. Without her insights and guidance, this research paper would not
have been possible.

To their previous research advisers, Mr. John Manuel Monteflor and Ms. Dharcee
Pelagio for their invaluable teachings, direction and recommendations in the study.

To Mr. Sherwin Malaga for his indispensable contribution in the statistical analysis of the
study’s raw data.

To the Bureau of Plant Industry for the verification of the Turmeric leaves, which is
significant in this study.

To Mr. Regie Perez for the assistance and guidance of conducting the experimentation
and data gathering at the Negros Prawn Producers Cooperative Analytical and Diagnostic
Laboratory.

To Mr. Kenneth Parra and Ms. Dharcee Pelagio for their assessment, corrections,
evaluations, and recommendations of their final research paper.

Lastly, the researchers would like to thank God for giving them the knowledge, strength,
and patience, and for his guidance throughout our study.
 TABLE OF CONTENTS

Page

TITLE PAGE I

APPROVAL SHEET II

ACKNOWLEDGEMENT III

TABLE OF CONTENTS IV

LIST OF TABLES VI

LIST OF FIGURES VII

ABSTRACT IX

INTRODUCTION 1

Background of the Study 1

Statement of the Problem 3

Hypothesis 4

Conceptual Framework 4

Scope and Limitations 6

Significance of the Study 6

Definition of Terms 7

Review of Related Literature 10

Properties of Turmeric 11

The Nature of Staphylococcus aureus 13

Antibacterial Efficacy of Ethanolic Extract 15

Ethanolic Extract against Staphylococcus aureus 17

Turmeric Leaf (Curcuma longa linn.) against Staphylococcus aureus 19

 Synthesis 20

MATERIALS AND METHODS 22

Research Design 22

Subjects of the Study 23

Materials and Equipments 23

Data Gathering Procedure 25

Pre-experiment 26

Experiment Proper 28

Post-Experiment 29

Statistical Treatment 30

Ethical Considerations 31

RESULTS AND DISCUSSION 33

Results 33

Discussion 35

CONCLUSION 37

RECOMMENDATIONS 39

REFERENCES 42

APPENDICES 47
 LIST OF TABLES

Table 1 Rating scale for p-value

31

Table 2 Antibacterial Activity using the zone of inhibition of Turmeric leaf

33

Table 3 Table for the Significant Difference in the zone of inhibition of 34

Turmeric leaves against Staphylococcus aureus using t-test.

 LIST OF FIGURES

Figure 1 Schematic Diagram of the research study’s variables 5

Figure 2 Flowchart of the data-gathering procedure 25

Figure 3 Plant certification of Turmeric leaves from Bureau of Plant Industry 47

Figure 4 Negros prawn document first page 47

Figure 5 Negros prawn document second page 47

Figure 6 Negros Prawn Request Analysis form 48

Figure 7 Raw data of Statistician results 48

Figure 8 Photo of researchers during the experiment 48

Figure 9 Experiment results 48

Figure 10 Experiment results 49

Figure 11 Photo of Turmeric leaves 50

Figure 12 Photo of Glass bottle 50

Figure 13 Petri dish 50

Figure 14 Staphylococcus aureus 50

Figure 15 Distilled water 50

Figure 16 Sterile swab 50

Figure 17 Forceps 51

Figure 18 Turmeric leaf ethanolic extract 51

Figure 19 Laboratory incubator 51

Figure 20 Extract using Maceration method 51

Figure 21 Turmeric leaf ethanolic extract 51

Figure 22 Preparation and sterilization of equipment and materials 51

Figure 23 Preparation of Staphylococcus aureus 52

Figure 24 Testing the concentration of extract against bacteria 52

Figure 25 Incubation of agar plates 52

Figure 26 Observation of zone of inhibition for negative control (Distilled water) 53

Figure 27 Data analysis for positive control (Gentamicin antibiotic) 53

Figure 28 Data analysis for 100% concentration of Ethanolic Extract 54

Figure 29 Disposal of laboratory waste using autoclave 54

 ABSTRACT

The rise of multi-drug resistant Staphylococcus aureus poses a significant global

healthcare challenge. Studies have highlighted the antibacterial properties of Turmeric
leaf, particularly curcumin, which can impede the replication of various viruses. This
research aimed to assess the antibacterial potential of Turmeric leaf ethanolic extract
against Staphylococcus aureus, alongside gentamicin and sterile distilled water. Extracts
were obtained through maceration in ethanol and applied to bacterial cultures, followed
by incubation at 37°C for 24 hours. Significant differences were observed in the zone of
inhibition, indicating Turmeric leaf's effectiveness against Staphylococcus aureus. The
data were analyzed using T-tests and mean, revealing a p-value of less than 0.001 and
rejecting the null hypothesis. Despite its lower efficacy compared to gentamicin,
Turmeric leaf extract showed promise as a natural alternative. Further investigation into
its antimicrobial mechanisms and optimization for clinical use is warranted. This study
lays groundwork for the development of natural antibacterial agents, offering potential
alternatives to synthetic antibiotics in the future.

Keywords: Antibacterial, Ethanolic Extract, Maceration, Staphylococcus aureus,

Turmeric leaf, Zone of inhibition
 1

INTRODUCTION

Background of the Study

Recent research has highlighted the potential antibacterial properties of natural

compounds, with turmeric leaves showing promise in studies for their antibacterial

effects. Meanwhile, antibiotic resistance is a growing global concern as it rendered

conventional antibiotics less effective in treating bacterial infections and necessitates the

exploration of alternative therapeutic strategies. The proposed research study, which

focused on the antibacterial activity of turmeric leaf ethanolic extract against

Staphylococcus aureus, held significant relevance in addressing several pressing issues

that are related to infectious diseases, healthcare, and the advancement of scientific

knowledge. As antibiotic resistance continues to grow, the urgent need for newly

developed ones becomes evident (Taylor & Unakal, 2022). This study’s exploration of

the potential antibacterial properties of a natural compound like the ethanolic extract of

turmeric leaf was poised to offer an alternative approach to countering Staphylococcus

aureus infections, potentially circumventing the limitations posed by the diminishing

efficacy of conventional synthetic antibiotics. In addition, Turmeric has a component

which is known as curcumin, and is the main reason why it can be used against different

bacteria, and it was also found out that it is safe to consume even at comparatively greater

quantities. It underwent substantial research as a nutraceutical ingredient for consumption

in functional foods due to its broad-spectrum biological activity and low toxicity

(Shirsath et al., 2017). There had been a growing interest when it comes to exploring the

potential antibacterial properties of natural compounds, and one such intriguing candidate
 2

for that is the Turmeric leaf, in which several studies had already reported that turmeric

leaves possess pharmacological properties, including antioxidant, anti-inflammatory,

antitumor, and antibacterial effects (Kim et al., 2021). Studies on curcumin's antibacterial

action are becoming more and more well-documented, showing that it has broad-

spectrum bacterial inhibitory characteristics (Moghadamtousi et al., 2014). A study was

able to determine that Staphylococcus aureus is a major human pathogen that causes a

wide range of infections. It is also a Gram-positive bacterium that is known for its

distinctive clustering and non-motile cocci shape and is a common inhabitant of the

human skin and nasal passages. This study offered valuable insights into the prevalence

and clinical implications of Staphylococcus aureus in diverse patient groups (Tong et al.,

2015).

While there were numerous studies conducted on combating the bacterium

Staphylococcus aureus on using potential alternative antibiotic medicines, there was still

insufficient research done locally about the potential antibacterial effects of Turmeric

leaf. Some researchers did not explore the potential of using Turmeric ethanolic extract in

combination with antibiotics or other natural products. Investigating synergistic effects

was crucial, specifically in the context of antibiotic resistance. Hence, this creates a gap

locally in research in the inclusivity of the Philippines in the involvement of finding

alternative medicine in combating Staphylococcus aureus. As a result, this could get the

attention of other researchers in gaining their interest to start researching about the

different plants in the Philippines that have potential antibacterial components.

In this study, the researchers identified the antibacterial activity of Turmeric

ethanolic extract against Staphylococcus aureus. The researchers hypothesized that there
 3

would be no significant difference in the zone of inhibition of Staphylococcus aureus

given varying concentrations of Turmeric leaf (Curcuma Longa Linn.) ethanolic extract.

In order to address the gap, the researchers conducted the study on antibacterial

properties of Turmeric leaf (Curcuma longa Linn.) ethanolic extract against

Staphylococcus aureus. Therefore, this study aimed to determine the potential of turmeric

leaf ethanolic extract as an antibacterial agent, which had the potential to directly benefit

individuals, communities, and healthcare systems by providing a new avenue in treating

Staphylococcus aureus infections, specifically in the Philippines.

Statement of the Problem

This study aimed to determine the effectiveness of turmeric leaf as an

antibacterial agent against Staphylococcus aureus.

Specifically, it aimed to answer the following questions:

1.) What is the effectiveness of turmeric leaf as an antibacterial agent using the

following concentration and controls:

a. 100 % Concentration

b. Positive Control - Gentamicin Antibiotic

2.) Is there a significant difference in the Zone of inhibition of Staphylococcus

aureus given the concentration of Turmeric leaf ethanolic extract and controls?

a. 100 % Concentration

b. Positive Control - Gentamicin Antibiotic

 4

Hypothesis

There is no significant difference in the zone of inhibition of Staphylococcus

aureus given the concentration of Turmeric leaf (Curcuma Longa Linn.) ethanolic extract

and its controls.

Conceptual Framework

The researchers aimed to experiment on the antibacterial activity of Turmeric leaf

(Curcuma longa Linn.) ethanolic extract against Staphylococcus aureus. It has the

potential to offer an alternative strategy for treating Staphylococcus aureus infections,

potentially getting around the constraints posed by the conventional synthetic antibiotics'

declining efficacy. As a result, this research was conducted in order to provide effective

treatment for Staphylococcus aureus infections. Furthermore, Ilham et al., (2018)

demonstrated that the turmeric leaf extract had substantial antibacterial activity, implying

that the turmeric leaf had a broad spectrum antimicrobial activity. Turmeric leaf extract

has a wide range of biological activities, including antibacterial, anti-inflammatory,

antimicrobial characteristics that can help cure Staphylococcus aureus infections.

 5

Independent Variable
Dependent Variable
Turmeric leaf (Curcuma longa Zone of Inhibition of S.
Linn) Ethanolic Extract (100%) aureus
Positive control - Gentamicin
Antibiotic

Figure 1. Conceptual Framework of the Study

The Schematic Diagram above illustrated the conceptual framework of the study

that exhibited the process by which the experiment was conducted. The figure included

(3) distinct variables: Independent, and Dependent. First, the researchers identified what

the independent variables were: the turmeric leaf, the concentration of the ethanolic

extract, and the positive and negative controls. These were the variables that were tested

and yielded results on the zone of inhibition of Staphylococcus aureus. In this study, the

zone of inhibition was the dependent variable. The change in this variable depended on

the manipulation of the independent variable. All of the aforementioned variables were

capable of directly and indirectly affecting the results of the experiment.

 6

Scopes and Limitations

This study focused on the antibacterial properties of Turmeric leaf (Curcuma

longa Linn.) ethanolic extract against Staphylococcus aureus. The study was conducted

from January 18, 2024 to January 22, 2024 during the second semester of the school year

of 2023-2024. The researchers tested the efficacy of Turmeric leaf (Curcuma longa

Linn.) ethanolic extract by culturing Staphylococcus aureus on agar plates and checking

for the zone of inhibition at different concentrations of Turmeric leaf (Curcuma longa

Linn.) ethanolic extract.

The other potentials of turmeric leaf were disregarded in this study, which only

concentrated on the antibacterial properties using the indicated amount of concentration.

This study only included Staphylococcus aureus as the dependent variable during the

duration of this study and this study only focused on Turmeric leaf (Curcuma longa

Linn.) ethanolic extract alone.

Significance of Study

The vital results of this study are highly significant and beneficial to the following:

Department of Health (DOH). This research serves as relevant information for

healthcare experts who are in the field of health sciences, particularly in antibacterial

studies and efficacy of the said variables. In which, they could use this as a substitute for

certain products that could be harmful to some extent.

 7

Department of Science and Technology (DOST). Turmeric extract is a promising

area of research for the development of new antibacterial agents. The Department of

Science and Technology may have invested in research into the antibacterial properties of

turmeric extract, particularly curcumin, to better understand how it could be used to fight

bacterial pathogens, including Staphylococcus aureus.

Pharmaceutical companies. This study may help pharmaceutical companies find

potential alternatives in treating diseases that have gained resistance to specific medicines.

Community. Due to inflation and low wages in the Philippines, some may have

encountered financial instabilities that did not allow them to purchase prescriptions and

medications for diseases and illnesses. Hence, this could give them insights about the uses

of the said plant that are common in the area to be used as an alternative medicine.

Future Researchers. This may serve as a review of related literature for upcoming

researchers for their research papers that are connected with bacteria and plant extracts to

widen their knowledge and improve their studies. These could also serve as a basis for

those who did not have a strong foundation for data analysis.

Definition of Terms

The following terms were defined conceptually and operationally to facilitate

better understanding of the study:

 8

Antibacterial. Anything that eradicates bacteria or inhibits their expansion or

reproduction. Antibacterial qualities can be found in heat, chemicals like chlorine, and

antibiotic medications. (Medical Definition of Antibacterial, 2021)

In this study, the antibacterial referred to substances that have the ability to kill or

inhibit the growth of bacteria. It is important to use antibacterial products appropriately

and only when necessary to help prevent the development of antibiotic-resistant bacteria

Bacteria. They are microscopic, single-celled organisms that thrive in diverse

environments. These organisms can live in soil, the ocean and inside the human gut.

Humans' relationship with bacteria is complex. Sometimes bacteria lend us a helping

hand, such as by curdling milk into yogurt or helping with our digestion. In other cases,

bacteria are destructive (Vidyasagar, 2019).

In this study, the bacteria being investigated was Staphylococcus aureus, which is

a gram-positive bacterium commonly associated with skin and tissue infections.

Ethanolic Extract. It is a process that is used in fine liquor distillation and is used

to filter out alcohol content from extracted material. The use of ethanol as an industrial

solvent is a cost-effective way to produce a quality extract from a large volume of plants.

Ethanol extraction systems use both cold and warm methods to extract easily through the

distillation process (Laboratory News from Lab Bulletin, 2021)

In this study, it was the extract that is being utilized in this study to sterilize or

prevent the Staphylococcus aureus bacteria from spreading.

 9

Maceration. Denotes the methodical immersion of a substance within a liquid

medium, usually a solvent, aimed at softening it to expedite the extraction of specific

compounds.

In this study, this process entailed submerging the material in the chosen solvent

for a predetermined duration, thereby enabling the selective extraction of target

constituents.

Staphylococcus aureus. It is a spherical bacteria commonly found on the skin and

mucous membranes of humans and animals. It can cause a range of infections, from

minor skin infections to severe and potentially life-threatening conditions.

In this study, Staphylococcus aureus was the main bacteria that was utilized to

prove the effectiveness of the Turmeric leaf ethanolic extract.

Turmeric (Curcuma longa Linn). Its rhizomes, which are native to Southwest

India, are the source of a bright yellow spice with a variety of medicinal properties. In the

tropics, it is widely cultivated and valued for its therapeutic properties. This article

compiles the therapeutic potentials of this plant as well as that of curcumin, one of its

bioactive components. (Benarba & Pandiella, 2018)

In this study, Turmeric was tested as a potential alternative medicine for bacteria

such as staphylococcus aureus and its effectiveness as an antimicrobial agent.

Zone of Inhibition. It is a circular region around antibacterial discs where

bacteria cannot flourish. This qualitative technique is used in clinical settings to assess
 10

antibiotic resistance and the ability of solids and textiles to prevent the growth of

microbes (Tanner, 2022).

In this study, it was used to determine the response of a bacteria to an antibiotic.

Review of Related Literature

Numerous studies have been conducted on combating the bacterium

Staphylococcus aureus using potential alternative medicines in the form of synthetic and

natural. One promising avenue of research involves investigating the antimicrobial

properties of oil extracts derived from various plants such as Turmeric. However, despite

the importance of the study, there is still insufficient research done locally about the

potential antimicrobial effects of the Turmeric leaf (Curcuma longa Linn.). Hence, this

creates a gap locally in research in the inclusivity of the Philippines in the involvement of

finding alternative medicine in combating the bacterium Staphylococcus aureus. To

address the gap, the researchers will conduct the study on antibacterial properties of

Turmeric leaf (Curcuma longa Linn.) ethanolic extract against Staphylococcus aureus.

Therefore, this study aims to determine the potential of turmeric leaf ethanolic extract as

an antibacterial agent, which has the potential to directly benefit individuals,

communities, and healthcare systems by providing a new avenue in treating

Staphylococcus aureus infections especially in the Philippines. The identified themes that

the literature review will focus on are the following: (1) Properties of Turmeric, (2) The

Nature of Staphylococcus aureus, (3) Antibacterial Efficacy of Ethanolic Extract, (4)

Ethanolic Extract against Staphylococcus aureus, and (5) Turmeric Leaf (Curcuma

Longa Linn.) against Staphylococcus aureus.

 11

Properties of Turmeric

The properties of turmeric encompass a wide range of health benefits, culinary

uses, and cultural significance, making it a versatile and valuable spice in various aspects

of life.

In an experimental study on knowing the antibacterial activities of different

turmeric extracts against Escherichia coli, Staphylococcus aureus, Salmonella typhi and

Candida albicans. This research investigates the antibacterial and preserving properties

of turmeric extracts in the food sector. Turmeric extracts in n-hexane, water, chloroform,

and ethanol were used as preservatives and antibacterial agents in meals.

Microbiologically (total bacterial, fungal, and coliform counts) and organoleptically

(color, odor, and taste) the samples were evaluated at day zero and at 15-day intervals.

Meals autoclaved for a shorter duration of time (5 minutes) and treated with a mixture of

1% or 2% turmeric extract were preserved for a longer period of time. Using the disk

diffusion method, Gul and Bakht (2013) determined the antibacterial activity of different

solvent extracted samples of turmeric. For disk diffusion assay, filter paper discs of 8

mm diameter were prepared and sterilized. Using sterile forceps, these discs were

aseptically placed over nutrient agar plates seeded with the respective test

microorganisms. Two different concentrations of turmeric extracts were aseptically

transferred to these discs. The plates were incubated in an upright position at 37 °C for 24

h. The diameters of inhibition zones (in mm) were measured. Afterwards, the results

showed that the turmeric extracts exhibited significant antibacterial activity against all the

tested bacteria.
 12

The study conducted by EL-Kenawy et al. in 2019 underscores the diverse array

of active ingredients that is present in turmeric, each contributing to a wide spectrum of

health benefits. In general, turmeric has been identified as a potent source of compounds

which exhibits anti-inflammatory, antimicrobial, antioxidant, antifungal, antibacterial,

antiviral, anti-ischemic, and antineoplastic activities. According to their own study, they

were also able to find out that the traditional uses of turmeric can span across a wide

range of health issues, which includes coughs, diabetes, dermatological issues, respiratory

difficulties, cardiovascular and hepatobiliary diseases, arthritis, irritable bowel syndrome

(IBS), peptic ulcers, psoriasis, and atherosclerosis. Even so, despite these therapeutic

potentials, their own study highlights certain challenges that can be associated with the

absorption and bioavailability of the active components that can be found in turmeric. It

is worth noting that these components face limitations in absorption, rapid digestion, and

swift excretion, which could potentially affect the efficacy as an antibacterial agent.

Ultimately, further research must be done in order to optimize and maximize the efficacy

and safety of turmeric based-intervention.

Furthermore, the study of Tyagi et al. (2015) demonstrated that the turmeric

extracts exhibited significant antibacterial activity against all the tested bacteria

(Escherichia coli, Staphylococcus aureus, Salmonella typhi) and the fungus Candida

albicans. The killing assay was performed as the procedure of the experiment in order to

test the effectiveness of turmeric extract. Bacterial cells were grown to the mid

logarithmic phase in BHI broth. The optical density (OD 600nm) of cells was adjusted to

0.5 (108 CFU/ml) in the 10 mM PBS buffer (pH 7.4) using a spectrophotometer. In the

presence of PBS, a final inoculum of 106 and 104 CFU/ml was subjected to different
 13

doses of curcumin I (25 M, 50 M, and 100 M). Aliquots were gathered at the specified

time periods (30, 60, and 120 minutes), placed on BHI agar plates in triplicate, and

incubated overnight at 37°C. The bacterial colonies were counted the next day, and

bacterial survival was represented as the mean percentage of survival vs the non-peptide

treated control (set at 100% survival). Each experiment was done three times on different

days. The findings, according to the researchers, show that curcumin I possesses strong

killing potential against all the tested bacteria. This suggests that turmeric extracts have

the potential to serve as effective antibacterial agents.

The aforementioned studies all share the same common findings regarding the

antibacterial activity of turmeric extracts, while differing in experimental methods and

the contextual information they provide about turmeric's medicinal properties. Together,

they underscore the potential of turmeric as a multifaceted natural remedy with promising

antibacterial properties.

The Nature of Staphylococcus aureus

Staphylococcus aureus, a versatile and formidable bacterium, presents a complex

and diverse nature with significant implications for human health and medical research.

According to Taylor and Unakal (2022) Staphylococcus aureus tends to be

grouped in clusters that are described as "grape-like." Between 18 and 40 degree Celsius,

these organisms can thrive facultatively or aerobically. Staphylococcus aureus may grow

in salt concentrations as high as 10%, and colonies tend to be golden or yellow (aureus

meaning golden or yellow).

 14

In their 2015 study on Staphylococcus infections, Tong et al., (2015) were able to

share their valuable insights on Staphylococcus aureus. They were able to determine that

it is a major human pathogen that causes a wide range of infections and it is also a Gram-

positive bacterium which is known for its distinctive clustering and non-motile cocci

shape, and is a common inhabitant of the human skin and nasal passages. This study

offers valuable insights into the prevalence and clinical implications of Staphylococcus

aureus in diverse patient groups. Such insights hold the potential to inform strategies for

both prevention and treatment in the realm of clinical practice, enhancing our

understanding of the complex relationship between this bacterium and human health.

In relation to this, Leung (2014) reported that Staphylococcus aureus is a

commensal organism that resides in skin and mucosa. Mild to life-threatening sepsis can

occur if the organism enters into the body especially in an immunocompromised or

immunosuppressed individual. Routes of entry include broken skin or mucosa, and oral

ingestion of infected food. Staphylococcus aureus can survive in a dry environment for

hours to months depending on the strain.

Around one third of healthy individuals carry this bacteria in their noses, pharynx

and on their skin. In a normal healthy and immunological person, Staphylococcus aureus

colonization of the skin, intestinal tract, or nasopharynx does not lead to any symptoms or

disease. (Ananya Mandal, 2019)

These studies share commonalities in providing information about

Staphylococcus aureus and its prevalence in humans, particularly in healthy individuals.

Although, the different studies vary in their specific focuses, with one emphasizing
 15

clinical implications, another discussing growth conditions, and highlighting prevalence

in healthy individuals. These differences reflect the diverse approaches of the researchers

in studying this bacterium and its relationship with human health.

Antibacterial Efficacy of Ethanolic Extract

The antibacterial efficacy of ethanolic extracts represents a critical area of study

in the field of research and medicine, offering potential solutions for combating bacterial

infections and exploring natural remedies.

According to a study conducted by Bitchagno et al. (2015) Crude ethanol extract

showed good activity against the bacteria strains tested with MIC of 64–256 μg/mL.

Among the isolated metabolites, 6-methyl-1,4-dihydroxyanthraquinone exhibited a strong

activity against Escherichia aerogenes with MIC of 16 μg/mL, while tectograndone

showed a moderate activity against Escherichia coli with MIC of 32 μg/mL.

The study of Valle et al. (2015) examined the leaf extracts of different Philippine

medicinal plants like guava (Psidium guajava), gale of the wind (Phyllanthus niruri),

fukien tea tree (Ehretia microphylla) and Piper betle (P. betle). The aforementioned

plants showed antibacterial activity against the gram-positive methicillin-resistant

Staphylococcus aureus and vancomycin-resistant Enterococcus. P. betle showed the

highest antibacterial activity for these bacteria in the disk diffusion (16–33 mm inhibition

diameter), minimum inhibitory concentration (19–156 μg/mL) and minimum bactericidal

concentration (312 μg/mL) assays. P. betle leaf extracts only showed remarkable

antibacterial activity for all the Gram-negative multidrug-resistant bacteria (extended

spectrum β-lactamase-producing, carbapenem-resistant Enterobacteriaceae and metallo-

 16

β-lactamase-producing) in the disk diffusion (17–21 mm inhibition diameter), minimum

inhibitory concentration (312–625 μg/mL) and minimum bactericidal concentration

(312–625 μg/mL) assays. They concluded that P. betle had the greatest potential value

against both Gram-negative and Gram-positive multidrug-resistant bacteria. Favorable

antagonistic activities were also exhibited by the ethanol extracts of Psidium guajava,

Phyllanthus niruri and Ehretia microphylla.

Moreover, the study of Altayb et al., (2022) sheds light on the promising

antibacterial properties of the methanolic extract of neem leaves. The findings of their

experiment indicate that the extract exhibited notable antibacterial activity against both

pathogenic and reference strains. Notably, the study also identifies Beta.d-

Mannofuranoside, O-geranyl as the most active compound within the extract of neem

leaves. This compound was not only able to demonstrate potent antibacterial activity but

it also satisfied all the five-rules of drug-likeness properties. Such adherence to drug-

likeness rules strengthens the compound's potential for further development. Lastly, this

could indicate a potential pathway for this compound to evolve into an alternative and a

commercially valuable antibacterial agent.

Bitchagno et al. (2015), Valle et al. (2015), and Altayb et al. (2022) decided to

investigate the antibacterial properties of natural plant extracts against different bacterial

strains that are commonly found around us such as Escherichia coli and Staphylococcus

[Link] similarities of the different studies includes how the researchers demonstrate

the antibacterial activity of the ethanolic extracts, the testing of specific bacterial strains,

and the use of Minimum Inhibitory Concentration (MIC) as a measure to determine the

potency of the ethanolic extracts to bacteria. However, the three studies vary in the
 17

methods of extraction of the different plant extracts. Bitchagno et al. (2015) used a crude

ethanol extract, Valle et al. (2015) examined leaf extracts from different medicinal plants,

and Altayb et al. (2022) focused on the methanolic extract of neem leaves.

Ethanolic Extract against Staphylococcus aureus

The investigation of ethanolic extract's effectiveness against Staphylococcus

aureus signifies a crucial exploration into potential remedies for combating this resilient

bacterium, with implications for both microbiological research and medical applications.

In perspective, the study, Antimicrobial Activity of Ethanol Extract of Rhizome

Turmeric (Curcuma Longa L.) For Growth of Escherichia coli, Staphylococcus aureus

and Candida albicans by Kasta (2020) showed that ethanolic extract of turmeric contains

alkaloids, flavonoids, saponins, tannins, and triterpenoid/steroid. The antimicrobial

inhibition of ethanolic extract of turmeric against Escherichia coli, Staphylococcus

aureus, and Candida albicans microbia at a concentration of 500mg/mL had a diameter

of 15.7 mm, 15 mm, 15.18 mm with a strong category. This confirms that turmeric could

be used as a possible novel antibacterial agent.

Moreover, a similar study conducted by Anjelina (2020) also concluded that the

ethanolic extract of kitolod leaves also shares the same properties of turmeric. It contains

alkaloids, flavonoids, and saponins. The antibacterial inhibition of ethanol extract of

kitolod leaves against Staphylococcus aureus and Salmonella typhi bacteria at a

concentration of 75% had a diameter of 11.3 mm and 12.16 mm with a strong category.
 18

In a study that was investigated by Salma et al., (2019), they were able to test the

antibacterial activity of ethanolic extract of cinnamon in different concentrations using

the disk diffusion method. The extracts of the experiment were prepared by using a

solvent ethanol. Staphylococcus aureus and Escherichia coli were tested for their activity

against a standard antibiotic Amikacin by disk diffusion method and the results were

compared with that of ethanolic extracts. As a result, the ethanolic extracts were more

effective against Staphylococcus aureus than Escherichia coli. Staphylococcus aureus

showed activity from 60% concentration and having a zone of inhibition of 17 mm while

Escherichia coli from 80% concentration and 18mm.

The three different studies are all about investigating the effectiveness of different

ethanolic extracts against the bacterium Staphylococcus aureus. Firstly, the findings of all

the researchers collectively suggests that ethanolic extracts from different plant sources

possess strong antibacterial properties because of phytochemicals that are present in the

various parts of plants. Secondly, the researchers also concluded that it could aid in

combating bacterial strains. However, the three studies vary in using different plant

sources, bacterial strains, and extract concentrations which result in different results of

the studies. Nevertheless, all of the three different studies support and encourage the

potential use of ethanolic extracts of plants as alternatives.

 19

Turmeric Leaf (Curcuma longa Linn.) against Staphylococcus aureus

Exploring the potential antibacterial properties of Turmeric Leaf (Curcuma longa

Linn.) against Staphylococcus aureus underscores a significant area of research with

promising implications for natural remedies and healthcare solutions.

In a comparative study conducted by Singh et al. (2017) which aimed to compare

the effectiveness of rhizome and leaf extracts from Turmeric (Curcuma longa Linn.) in

inhibiting the growth of pathogenic bacteria, showed that both the turmeric leaf and the

turmeric rhizome contains antibacterial properties, but their effects were only specific to

certain bacterial strains. Bacterial strains such as Klebsiella pneumoniae, Staphylococcus

aureus, Pseudomonas aeruginosa and Bacillus subtilis were tested. Notably,

Pseudomonas aeruginosa and Bacillus subtilis were significantly inhibited by both

extracts. After they compared the antimicrobial activity of the two extracts, it showed that

the rhizome extract exhibited greater potency than the leaf extract against the tested

bacteria.

In a study by Ilham et al., (2018) they were able to discover that the bioactive

chemicals in the extract also gained with concentration, which boosted the diffusion rate

of antimicrobial substances and led to a bigger diameter of inhibitory zones. The resulting

inhibitory zones responded differently to various concentrations. The ability of the

antibacterial agent to penetrate the agar plate and the sensitivity of the bacteria to the

antibacterial substance both had an impact on the diameter of the inhibitory zone.

Additionally, the results of their experiment showed that the extracts of turmeric leaves

dissolved in ethyl acetate contained alkaloid, flavonoid, glycoside, saponin, and

 20

triterpenoid/steroid compounds, while those that were dissolved in in methanol solvents

contained alkaloids, flavonoids, glycosides and saponins. Lastly, they were able to

conclude that Turmeric leaf extract is an effective antibacterial agent and also it was one

of the good media for the growth of a good bacteria which is called Lactobacillus

acidophilus bacteria.

These studies all discussed how turmeric leaf affects Staphylococcus aureus based

on various tests that have uncovered the substance's novel characteristics and

antibacterial activity. Researchers were able to identify distinct characteristics that

promoted the development of inhibitory zones as well as effects that were present

exclusively in particular bacterial strains.

Synthesis

In general, several studies investigated the effectiveness of ethanolic extracts

against Staphylococcus aureus, a bacterium with significant implications for

microbiological research and medical applications. The findings suggested that ethanolic

extracts from various plant sources had strong antibacterial properties due to

phytochemicals present in plants. These extracts could aid in combating bacterial strains.

However, the studies varied in their use of plant sources, bacterial strains, and extract

concentrations, resulting in different results. Turmeric leaf, a key component of turmeric,

was identified for its unique antibacterial properties and inhibitory zones.

The proposed research study, which focused on the antibacterial activity of

turmeric leaf ethanolic extract against Staphylococcus aureus, held significant relevance

in addressing several pressing issues related to infectious diseases, healthcare, and the
 21

advancement of scientific knowledge. As antibiotic resistance continued to grow, the

urgent need for newly developed ones became evident (Taylor & Unakal, 2022).

In addition, this study's exploration of the potential antibacterial properties of a

natural compound like the ethanolic extract of turmeric leaf was poised to offer an

alternative approach to countering Staphylococcus aureus infections, potentially

circumventing the limitations posed by the diminishing efficacy of conventional synthetic

antibiotics. Furthermore, this study was highly important due to the increasing number of

multidrug-resistant Staphylococcus aureus, including Methicillin-resistant

Staphylococcus aureus (MRSA), which elevated the importance of finding innovative

solutions to tackle these infections.

The present study could be beneficial as it would serve as relevant information for

healthcare experts in the field of health sciences with their antibacterial studies and

efficacy of the said variables. The ethanolic extract of turmeric leaf would be utilized as a

means of substitute for certain medicines that could be harmful to some extent. In

addition, it would also contribute to the scope and body of knowledge, broadening the

knowledge on antibiotic resistance in the Philippines. Considering the gap of lack of

information and local studies regarding turmeric leaf as a potential alternative medicine

for bacteria, the researchers conducted the study to fill in the literature gaps. Therefore,

the researchers presented newly revised knowledge and insights that would enlighten the

beneficiaries.
 22

MATERIALS AND METHODS

This section presents the research design, subjects of the study, materials and

equipment, data-gathering procedure, statistical treatment, and the ethical considerations.

Research Design

An experimental research approach was employed to address the questions posed

in this study. The experimental method is defined by McLeod (2023) as “The

manipulation of variables in order to establish cause-and-effect relationships.” This

particular research method was appropriate for this study since it essentially required the

researchers to control the different concentrations of the ethanolic extract of turmeric leaf

and identify their impacts on Staphylococcus aureus. This research design made use of t-

test and mean as the statistical tools that determined the effect of the independent variable

on the dependent variable and proved whether turmeric leaf is effective against

Staphylococcus aureus. Hence, the researchers performed the experiment in a laboratory

and utilized the Kirby-Bauer disk diffusion method, which was defined by Christenson et

al. (2017) as a standardized technique for testing rapidly growing pathogens. Considering

that one of the key objectives of this study was to determine the effectiveness of the

turmeric leaf extract on the bacteria, this research design is suitable for this study.
 23

Subjects of the Study

In this research study, the primary subjects that were investigated were

Staphylococcus aureus and Turmeric leaf (Curcuma longa Linn) ethanolic extract. The

100% concentration of turmeric leaf ethanolic extract and the positive and negative

controls was the independent variable, and the zone of inhibition of Staphylococcus

aureus was the dependent variable.

Materials and Equipment

The following materials were utilized in this study:

● Seven (7) pairs of Disposable gloves

● Seven (7) Lab coats

● Five-hundred (500) grams of Turmeric leaves

● One (1) Laboratory oven

● One (1) Rotary evaporator

● Thirty-eight (38) grams of Mueller Hinton agar powder

● One thousand two-hundred fifty (1250) milliliters of Sterile distilled water

● Thirty (30) Petri dishes

● One (1) Autoclave

● One (1) Laboratory refrigerator

● One (1) culture of Staphylococcus aureus

● Ninety-nine and five tenths (99.5) milliliter of One (1) percent of Sulfuric acid
 24

● Zero and five tenths (0.5) milliliters of One and one hundred seventy-five

thousandths (1.175%) BaCl2·2H2O

● One (1) Spectrophotometer

● Thirty (30) Sterile swabs

● Three (3) Beakers

● One (1) Forceps

● One (1) Laminar flow hood

● One (1) Laboratory incubator

● One (1) Caliper

 25

Data-Gathering Procedure

PRE-EXPERIMENT

Preparation of Turmeric

Extraction of Turmeric

Preparation of Mueller-

Collection of

Sterilization of

EXPERIMENT PROPER

Preparation of

Testing the concentration of

Turmeric leaf ethanolic extract
against Staphylococcus aureus
and its controls

Incubation of Agar

POST EXPERIMENT

Observation of Zone of

Data Analysis

Disposal of Laboratory
 26

Figure 2. Flowchart of the Data Gathering Procedure

Pre-Experiment Phase

The study was conducted in the laboratory of Negros Prawn Producers Cooperative

Analytical and Diagnostic Laboratory, Bacolod City, Negros Occidental. The laboratory

was handled by trained professionals and was safe and well-equipped which was utilized

by the researchers for guidance and their experimental study. Furthermore, the results

were determined based on the antibacterial activity of Turmeric leaf (Curcuma longa

Linn.) ethanolic extract with a basis to the (+) Gentamicin antibiotic and (-) sterile

distilled water when used against Staphylococcus aureus through the measurement of the

zone of inhibition.

I. Preparation of Turmeric leaves (Curcuma longa linn.)

The researchers collected 500 grams of Turmeric leaves, which were used as the

independent variable in the study. The Turmeric leaves were purchased locally from a

local farm in the city of Candoni. To ensure that there were no problems, the researchers

sent a sample to the Bureau of Plant Industry and received a certificate of authenticity

that ensured the classification and quality of the Turmeric leaves. Then the leaves were

delivered to the Negros Prawn Producers Cooperative Analytical and Diagnostic

Laboratory for the experimentation process.

II. Extraction of Turmeric leaves (Curcuma longa linn.)

The leaf extract was obtained through the Maceration method, a process that

involved a solvent (70% ethanol). Going through the preparation process, the person-in-
 27

charge from the Negros Prawn Producers Cooperative Analytical and Diagnostic

Laboratory dried the Turmeric leaves using a thermal oven at 45 degrees Celsius. After

drying, it was then crushed into tiny pieces, placed into the rotary flask with the solvent,

and agitated for better extraction. Then the crushed leaves were placed in a rotary

evaporator. The rotary evaporator rotated at a rate of 60 rotations per minute and

consisted of a cooler with a temperature of 9°C for the condenser and a heating bath for

the rotary flask with a temperature reaching 70°C. The mixture remained in the rotary

evaporator until the dripping stopped for at least 30 seconds.

III. Preparation of Mueller-Hinton agar plates

The researchers started by weighing 38 grams of Mueller-Hinton agar powder and

then mixing it with 1000 milliliter of distilled water while ensuring even dispersion

through gentle stirring. They then sterilized it using an autoclave at 121°C for 15

minutes. The liquid agar was then poured into sterile petri dishes, cooled until solidified,

labeled with essential information, and stored in a refrigerator for later use in microbial

analysis linked to ethanol extraction.

IV. Collection of Staphylococcus aureus

The bacterium sample of Staphylococcus aureus was purchased and provided by

the Negros Prawn Producers Cooperative for the study.

V. Sterilization of Equipment

The Negros Prawn Producers Cooperative Laboratory thoroughly prepared the

materials and equipment that was used for the experiment. To ensure that the
 28

environment was safe, clean, and free of pollutants, all equipment utilized in the

experiment proper were dusted.

Experiment Proper (Kirby-Bauer disk diffusion)

I. Preparation of Staphylococcus aureus

The experiment proper began with the cultivation of a pure Staphylococcus

aureus culture. Then, the Negros Prawn Producers Cooperative Analytical and

Diagnostic Laboratory prepared the McFarland's Standard. 100 mL of McFarland's

standard was prepared by mixing 99.5 mL of 1% sulfuric acid and 0.5 mL of 1.175%

8
BaCl2·2H2O. The turbidity was equivalent to 1.0 x 10 CFU/ml bacteria. Bacterial

isolates from the pure culture of Staphylococcus aureus were then transferred to a sterile

buffered phosphate or distilled water solution, and the turbidity was adjusted using a UV-

visible spectrophotometer at 600 nm wavelength in comparison with the absorbance of

the McFarland's standard, 0.089.

II. Testing the concentration of Turmeric leaf ethanolic extract against

Staphylococcus aureus and its controls

The sterile swab per organism was dipped into the broth culture of the organism

and gently squeezed against the inside of the tube to remove excess fluid from the swab.

The swab with the test organism was streaked evenly on each of the sterile Mueller-

Hinton agar (MHA) plates. After the streaking was completed, the plates were given time

to dry for 5 minutes. Then, the discs (from the 100% Turmeric leaf ethanolic extract, (+)

control Gentamicin antibiotic, and the (-) control distilled water) were placed on the
 29

surface of each agar using sterilized forceps. The discs were gently pressed into the

surface of the agar using flame-sterilized forceps or an inoculation loop.

III. Incubation of Agar Plates

The plates were then carefully inverted and incubated for 24 hours at 37°C. After

incubation, a caliper was used to measure the diameter of the zone of inhibition for each

of the discs used.

Post-experiment

I. Observation of Zone of Inhibition

Following the application of Turmeric leaf (Curcumin longa linn.) ethanolic

extract using the Kirby-Bauer disk diffusion method, the plates were carefully observed.

A caliper was also used in measuring the zone of inhibition of the independent variables

(from the 100% Turmeric leaf (Curcumin longa linn.) ethanolic extract, (+) Gentamicin

antibiotic, and (-) distilled water) against Staphylococcus aureus. This observation was a

critical step in assessing the antibacterial activity of the extract against Staphylococcus

aureus.

II. Data Analysis

Afterwards, the results that were gathered from the procedures of the experiment

were arranged and analyzed thoroughly by using the statistical tools (Mean and Analysis

of Variance) to garner information in answering the statement of the problem. The

 30

researchers were also able to make sure that all steps and procedures of the experiment

had been followed thoroughly.

III. Disposal of Laboratory Waste

After the experiment was conducted, the researchers and the person in charge

were responsible for returning the materials and equipment in their proper places and also

in disposing of the bacteria properly in order to avoid contamination.

Statistical Treatment

In order to answer the research questions posed in this study, the following

statistical tools were used.

To answer the problem statement 1: What is the effectiveness of turmeric leaf as

an antibacterial agent using the following variables: (a) 100 % concentration, (b) positive

control - gentamicin, (c) negative control - distilled water, mean will be used to

determine the level of effectiveness of the treatments, as determined by the zone of

inhibition of the Staphylococcus aureus.

To answer the problem statement 2: Is there a significant difference in the zone

of inhibition of Staphylococcus aureus given the varying variables of turmeric leaf

ethanolic extract? (a) 100 % concentration, (b) positive control - gentamicin antibiotic,

(c) negative control - distilled water, and t-test will be used to compare the means of the

zone of inhibition across the different variables of the study.

 31

Furthermore, this study will be utilizing 0.05 as the basis for the significance

level. Hence, the rating scale below will be used to interpret the p-value to determine the

significance:

Table 1

p - value Interpretation

p > 0.05 No Significant Difference

p < 0.05 Significant Difference

Ethical Consideration

This experimental study was carefully designed to uphold ethical principles and

ensure the responsible conduct of scientific inquiry. The researchers were deeply

committed to mitigating any potential environmental impact resulting from the use of

Turmeric leaf ethanolic extract. Precautions were taken to prevent harm to the

environment, including the proper disposal of chemical waste, to prevent any

contamination of natural ecosystems. The researchers were also dedicated to adhering to

best practices in laboratory research. Safety measures were rigorously followed to ensure

the health and well-being of all individuals involved in this research experiment.

Specifically, in handling the bacteria, strict protocols for containment, handling, and

disposal were implemented to minimize any potential risks and uphold the highest

standards of biosafety. Thus, the researchers also ensured that the experiment avoided

any unethical behavior such as fabricating results after the experiment was conducted.
 32

RESULTS AND DISCUSSION

 33

This section of the research paper presents the results, analysis, and discussion of

the gathered data. Results are presented in the following tables containing specific details

and followed by a corresponding comprehensive discussion to satisfy the necessity of

answering the statement of the problem and the formulated hypothesis. This study

contains three treatments, with ten trials each. Namely, (a) Gentamicin antibiotic, (b)

100% Turmeric leaf ethanolic extract, and (c) distilled water.

Table 2

Antibacterial Activity using the zone of inhibition of the different treatments against

Staphylococcus aureus.

Treatments Mean SD Interpretation

Gentamicin Antibiotic 34.1 3.31 Most Effective

100 % Turmeric Leaf

21.2 1.81 Effective
Ethanolic Extract

Distilled Water 0 0 Least Effective

Table 2 illustrates the results of the following treatments involving Gentamicin

Antibiotic, 100% Turmeric leaf Ethanolic Extract, and Distilled water. The Gentamicin

Antibiotic had a promising result of a mean of 34.1 using the zone of inhibition, implying

that it is highly effective. On the other hand, the 100% Turmeric leaf Ethanolic Extract

had a mean of 21.2, which indicates that its effectiveness is normal. Finally, the mean of

Distilled water exhibited unsatisfactory results, registering a value of zero and proving it

to be ineffective.
 34

Table 3

Test for the Significant Difference in the zone of inhibition of Turmeric leaves against

Staphylococcus aureus using t-test.

F df p Interpretation

With

Zone of Inhibition -10.8 18 <.001 Significant

Difference

Table 3 illustrates the results of the t-test conducted to evaluate the significant

difference in the zone of inhibition of Staphylococcus aureus with the use of turmeric leaf

extract. With a calculated F-value of -10.8, p-value that is less than .001 and with the

degrees of freedom (df) of 18, indicating that there is a significant difference in the

antibacterial activity among the different treatments. Furthermore, the data also suggests

that there is a notable difference in the antibacterial activity among the different

treatments. Despite the fact that the negative F-value implies an inverse relationship

between the variables, the statistical significance suggests that this difference is unlikely

due to chance. This aligns with the existing studies that highlights the potent antibacterial

properties of turmeric and its active compound, curcumin, against Staphylococcus

aureus. According to a study of Goyad et al. (2015), turmeric leaf has also been

investigated for its antibacterial potential. Turmeric leaf extract demonstrated notable

antibacterial activity against Staphylococcus aureus, among other pathogens.

Discussion
 35

Turmeric Leaf Effectiveness Evaluation

The main contributor to the effectiveness of the Turmeric leaf is the presence of its

bioactive compound which is curcumin. Curcumin, an important constituent of turmeric

that helps in treating bacterial infections. Many studies showed that curcumin exhibited

antibacterial activities against Gram-negative and Gram-positive bacteria. The

antibacterial action of curcumin involves the disruption of the bacterial membrane,

inhibition of the production of bacterial virulence factors and biofilm formation, and the

induction of oxidative stress. These characteristics also contribute to explain how

curcumin acts as a broad-spectrum antibacterial adjuvant, which was evidenced by the

markedly additive or synergistic effects with various types of conventional antibiotics or

non-antibiotic compounds (Dai et al., 2022).

Regarding the assessment of the effectiveness against Staphylococcus aureus, the

findings from Table 2 offer valuable insights into the various performance of the

different treatments. The results indicate the efficacy of different interventions based on

their respective mean outcomes.

Gentamicin Antibiotic, as depicted in Table 2, emerges as a standout performer

with a substantial mean zone of inhibition of 34.1 This result suggests a high level of

effectiveness in inhibiting the growth or presence of Staphylococcus aureus, positioning

Gentamicin Antibiotic as a highly effective solution for combating this pathogen.

In contrast, the 100% Turmeric leaf Ethanolic Extract demonstrates a lower mean

value of 21.2. While still effective, this result indicates a modern level of efficacy

compared to Gentamicin Antibiotic. It suggests that Turmeric leaf extract can serve as a
 36

reasonably effective intervention against Staphylococcus aureus, although with a slightly

reduced potency.

Conversely, the mean value of Distilled water is zero, indicating that there was no

observable effect of the zone of inhibition against Staphylococcus aureus. This result

reaffirms the ineffectiveness of Distilled water as a control agent for this pathogen,

providing a baseline for comparison against active treatments.

 37

CONCLUSION

Recent research has spotlighted the potential antibacterial properties of natural

compounds, with turmeric leaves emerging as a promising candidate due to their reported

antibacterial effects. This investigation was prompted by the escalating global concern of

antibiotic resistance, which rendered conventional antibiotics less effective in combating

bacterial infections, necessitating the exploration of alternative therapeutic strategies.

Moreover, the well-documented antibacterial action of curcumin against a wide range of

bacteria further underscores its potential as an alternative therapeutic agent. This study

aimed to investigate the antibacterial activity of turmeric leaf ethanolic extract against

Staphylococcus aureus. The researchers hypothesized that the turmeric leaf ethanolic

extract would not significantly alter the zone of inhibition against Staphylococcus aureus.

By addressing this research gap, the study sought to determine the potential of turmeric

leaf ethanolic extract as an antibacterial agent, with implications for enhancing treatment

options for Staphylococcus aureus infections, particularly in the Philippines.

The investigation revealed promising results regarding the antibacterial activity of

turmeric leaf ethanolic extract against Staphylococcus aureus. The extract exhibited a

substantial zone of inhibition, indicative of its efficacy as an alternative antibacterial

agent. These findings support the hypothesis that varying concentrations of turmeric leaf

ethanolic extract maintain consistent antibacterial properties against Staphylococcus

aureus, thereby validating its potential as a natural remedy for combating bacterial

infections.
 38

In conclusion, our study investigated the potential antibacterial activity of an

ethanolic extract from turmeric leaves against Staphylococcus aureus, a bacteria of

significant clinical concern due to its resistance to antibiotics. Our findings support the

notion that turmeric leaf extract possesses antibacterial properties, as evidenced by the

significant zone of inhibition observed in our experiments.

However, it's important to note that our study only utilized a single concentration

(100%) of the turmeric leaf extract. Future research should explore a range of

concentrations to establish dose-response relationships and determine the optimal

concentration for maximal antibacterial efficacy. Additionally, further investigations

could delve into the mechanisms underlying the antibacterial activity of turmeric leaf

extract and its potential synergistic effects with other antibacterial agents.

Despite these limitations, our study underscores the potential of turmeric leaf

extract as a natural alternative for combating Staphylococcus aureus infections,

contributing to the ongoing efforts to address antibiotic resistance. Continued research in

this area holds promise for the development of novel therapeutic strategies against

bacterial infections.
 39

Recommendations

Given the significant potential demonstrated by the research on the antibacterial

properties of Turmeric leaf ethanolic extract against Staphylococcus aureus, it is highly

recommended that The Department of Science and Technology (DOST) invest further

in this area of study. Turmeric extract, particularly curcumin, presents itself as a

promising avenue for the development of novel antibacterial agents. By supporting

research initiatives in this field, the DOST can contribute to the advancement of

knowledge regarding the efficacy and mechanisms of action of turmeric extract against

bacterial pathogens, including Staphylococcus aureus.

Such investments hold the promise of yielding innovative solutions for combating

infectious diseases, thereby benefiting individuals, communities, and healthcare systems,

particularly in the context of the Philippines where Staphylococcus aureus infections

pose a significant health concern. By fostering research and development in this

direction, the DOST can potentially facilitate the translation of scientific discoveries into

practical applications that improve public health outcomes and address pressing

healthcare challenges in the nation.

Future researchers interested in exploring the antibacterial properties of plant

extracts can build upon the findings of this study to deepen our understanding and

potentially develop new avenues for combating bacterial infections. To further advance

this field of research, it is recommended to conduct comprehensive investigations into the

specific mechanisms of action underlying the antibacterial activity observed in Turmeric

 40

leaf extract. This could involve elucidating the active compounds responsible for the

observed effects and their interactions with bacterial cells.

Additionally, exploring the potential synergistic effects of combining Turmeric

leaf extract with other antimicrobial agents could enhance its efficacy against

Staphylococcus aureus and possibly other bacterial pathogens. Furthermore, future

studies could focus on optimizing extraction methods to maximize the yield of bioactive

compounds from Turmeric leaves while ensuring sustainability and minimal

environmental impact.

By addressing these aspects, future research endeavors can contribute to the

development of novel therapeutic strategies for combating bacterial infections, thereby

benefiting individuals, communities, and healthcare systems, particularly in regions like

the Philippines where such infections pose significant public health challenges.

For the food industry, particularly in food preservation and safety, understanding

the antibacterial properties of Turmeric leaf extract could offer insights into its potential

use as a natural preservative, enhancing food shelf life and reducing the need for

chemical additives.

Research into the antibacterial effects of turmeric leaf extract could help

determine its efficacy against common foodborne pathogens, potentially extending shelf

life while minimizing reliance on synthetic preservatives. This exploration aligns with the

growing consumer demand for natural, minimally processed foods, offering both safety

and sustainability benefits for the food industry.

 41

Incorporating findings from research on natural remedies and antimicrobial

resistance into curriculum materials for the Department of Health (DOH) presents a

multifaceted opportunity. Firstly, it enables students to grasp the significance of

traditional remedies in combating diseases, offering a nuanced understanding beyond

conventional pharmaceutical interventions.

Furthermore, integrating this research into the curriculum promotes

interdisciplinary learning, as students explore connections between biology, chemistry,

and health sciences. They can analyze how various natural compounds interact with

microbial pathogens on a molecular level, deepening their comprehension of both

biological processes and chemical properties.

Additionally, studying antimicrobial resistance within the context of natural

remedies encourages students to consider broader societal implications, such as the

importance of conservation and sustainable use of medicinal plants.

Overall, this approach not only enhances students' academic knowledge but also

cultivates a holistic perspective on healthcare, empowering them to approach future

health challenges with a comprehensive understanding of both traditional and modern

treatment modalities
 42

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APPENDICES

Appendix A
Photos and documents

Figure 3. BPI document Figure 4. Negros Prawn Figure 5. Negros Prawn

document first page document second page
 48

Figure 8. Photo of researchers

during the experiment

Figure 7. Raw data of

Statistician results

Figure 6. Negros Prawn

Request Analysis form

Figure 9. Experiment results

 49

Figure 10. Experiment results

 50

Appendix B
Materials

Figure 11. Turmeric leaves Figure 12. Glass Bottle

Figure 13. Petri dish

Figure
Figure 14. Staphylococcus Figure 15. Distilled water
16. Sterile swab
aureus
 51

Figure 17. Forceps

Figure 18. Turmeric leaf
ethanolic extract Figu
re 19. Laboratory incubator

Appendix C
Pre-experiment: Ethanolic extraction of Turmeric leaves

Figure 20. Extract using Figure 21. Turmeric leaf Figure 22. Preparation and
Maceration method ethanolic extract sterilization of equipment and
materials

Appendix D
 52

Experiment proper

Figure 23. Preparation of Figure 24. Testing the Figure 25. Incubation of agar
Staphylococcus aureus concentration of extract plates
against bacteria

Appendix E
 53

Post-experiment

Figure 26. Observation of Zone of inhibition for negative control (Distilled water)

Figure 27. Data analysis for positive control (Gentamicin antibiotic)

Figure 28. Data analysis for 100% concentration of Ethanolic extract

 54

Figure 29. Disposal of laboratory waste using autoclave