FAR EASTERN UNIVERSITY

HIGH SCHOOL

“The Antibacterial Activity of Tinospora cordifolia (Makabuhay Plant) against Staphylococcus

aureus”

A Research Paper Presented to

Science, Technology, Engineering and Mathematics (STEM)
Far Eastern University High School

In Partial Fulfillment
of the Requirements in Practical Research 2

by:

Baricaua, Mayflor B.
Baydo, Nyzel Jheune Z.
De Guzman, Alcyren Joy D
De Guzman, Giselle Jane C.
Enciso, Slinnie C.
Landrito, Sofia Lubelle P.
Lorio, Alliah B.
Nituda, Jasmine Claire M.
Santos, Mernelle Mae O.

Grade 12 – STEM 2

October 2018

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 APPROVAL SHEET

In partial fulfillment of the course requirements in Practical Research 2, this research paper has been
prepared and presented by the undersigned members of Grade 12 STEM 2, and is hereby requested
for corresponding approval and acceptance of this requirement.

_______________________ _______________________
Mayflor B. Baricaua Sofia Lubelle P. Landrito

_______________________ _______________________
Nyzel Jheune Z. Baydo Alliah B. Lorio

_______________________ _______________________
Alcyren Joy D. De Guzman Jasmine Claire M. Nituda

_______________________ _______________________
Giselle Jane C. De Guzman Mernelle Mae O. Santos

_______________________
Slinnie C. Enciso

Approved and accepted in partial fulfillment of the course requirements in Practical Research 2 under
the supervision of Science, Technology, Engineering, and Mathematics (STEM) Department of Far
Eastern University High School.

_____________________
Mr. Glen S. Mirabete
Research Adviser

Date: ________________

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 ACKNOWLEDGMENT

We want to express our deepest gratitude to our one and only teacher in Practical Research 2,

Mr. Glen Mirabete. Without him, it would not be possible for us to accomplish and finish this research

paper on time. We would also like to thank our marvelous institute, Far Eastern University High School,

for always giving us the opportunity to show our potentials.

We would also like to give our deepest appreciation to Ms. Aza Freya Lan A. Tubato for being

our research assistant and we would also like to thank the biology department of Adamson Univers ity.

We would also like to extend our deepest appreciation to our friends for supporting us and for

all the encouragement they have given us. More importantly to our parents who supported us not just

morally but also financially from the very beginning until this day.

And above all, to the origin of all wisdom. We would like to express our appreciation to our

Almighty God for giving us the strength and wisdom. To God be all the glory!

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 TABLE OF CONTENTS

Title Page i
Approval Sheet ii
Acknowledgment iii
Table of Contents iv
List of Tables v
List of Figures v
Abstract vi

1.0 The Problem and Its Background

1.1 Background of the Study 1
1.2 Theoretical Framework 2
1.3 Statement of the Problem 3
1.4 Research Hypotheses 4
1.5 Significance of the Study 5
1.6 Scope and Delimitations 6
1.7 Definition of Terms 7

2.0 Review of Related Literature and Studies

2.1 Biological and Phytochemical Analysis of Tinospora cordifolia 18
2.2 Antibacterial Activity of Tinospora cordifolia 19
2.3 Staphylococcus aureus 21
2.4 Synthesis of Related Literature and Studies 22
2.5 Conceptual Framework 23

3.0 Research Methodology

3.1 Research Design 25
3.2 Materials and Study Site 26
3.3 Data Collecting Instruments 26
3.4 Data Gathering Procedure 28
3.5 Data Analysis 31
3.6 Ethical Considerations 31

4.0 Presentation, Analysis, and Interpretation of Data

4.1 Antibacterial Properties of Tinospora cordifolia 32
4.2 Zone of Inhibitions 33
4.3 Analysis of Variance (ANOVA) 35

5.0 Summary of Findings, Conclusions, and Recommendations

5.1 Summary of Findings 39
5.2 Conclusions 40
5.3 Recommendations 40

References 42

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Appendices
Appendix A: Permit to Conduct Research 47
Appendix B: RRL Summary 48
Appendix C: Documentation 74
Appendix D: Statistical Summary 77
Appendix E: Phytochemical Analysis 81
Appendix F: Breakdown of Expenses 82

Curriculum Vitae 83

LIST OF TABLES

Table 3.3 Instruments 26

Table 3.4 Methodology Flowchart 31
Table 4.1 Phytochemical Analysis of the Ethanolic extract 32
of Tinospora cordifolia
Table 4.2.1 Guideline chart for susceptibility 33
Table 4.2.2 Zone of inhibition of the negative control 33
Table 4.2.3 Zone of inhibition of the positive control 34
Table 4.2.2 Zone of inhibition of the different concentrations 35
Table 4.3.1 Summary of the count, sum, average and variance 35
of the concentrations
Table 4.3.2 Summary of the count, sum, average and variance 35
of the distilled water
Table 4.3.3 Summary of the count, sum, average and variance 36
of the Clindamycin
Table 4.3.4 Descriptive Statistics 36
Table 4.3.5 Estimated Marginal Means 37
Table 4.3.6 Analysis of Variance 38

LIST OF FIGURES

Figure 1.2 Theoretical Framework of the Study 3

Figure 2.1 Conceptual Framework of the Study 24

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 ABSTRACT

This research was conducted to know the antibacterial activity of Tinospora cordifolia against

Staphylococcus aureus. The researchers aim to answer three problems which were; What are the

antibacterial properties of Tinospora cordifolia?, Does the ethanolic extract of Tinospora cordifolia

stem in 50%, 75%, and 100% concentrations against Staphylococcus aureus capable of producing zone

of inhibition using Kirby-Bauer method?, and lastly, Is there a significant difference between the 50%,

75%, and 100% concentrations compared to Clindamycin in inhibiting Staphylococcus aureus? All the

three were answered in this study using experimentation. The data analysis that were used is ANOVA

or Analysis of Variance. The ethanolic extract was brought to Adamson University for phytochemica l

testing. Results showed that the ethanolic extract of the T. cordifolia stem contains flavono ids,

glycosides, tannins, amino acids and alkaloids which are a bioactive component of a plant. 100%, 75%

and 50% are the concentrations that were used in this study. The positive control is the clindamyc in

and the negative control is the distilled water. Antibacterial Susceptibility Test or Kirby-Bauer Method

was used to test the susceptibility of the Staphylococcus aureus. Results showed that there is a

significant difference between the zone of inhibitions produced by the 100% ethanolic extract of T.

cordifolia and the 75%,50% ethanolic extract of T. cordifolia and distilled water compared to

clindamycin in inhibiting the growth of Staphylococcus aureus.

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 Chapter 1

THE PROBLEM AND ITS BACKGROUND

This chapter presents the introduction of the study, what the study is all about and where it came

from. It also included the statement of the problem, theoretical framework, significance, scope and

delimitation, and the definition of terms.

1.1 Background of the Study

Herbal medicines have been part of traditional health care in most parts of the world for

thousands of years. Back in the day when there are little or few resources and when some doesn’t

have the means to obtain the things they need, people learned to study their surroundings and

take full advantage of what is around them that can and will benefit them. One example is when

people back then needs to be treated because of some disease, illness or an injury they rely on

the available resources that they can get especially around the place they live in and that was

when they discovered the use of some plants as herbal medicines or as an alternative medicine.

One of the plants known for having many medicinal use in traditional system of

medicine is the Tinospora cordifolia. It is a common climbing shrub found in tropical deciduous

forest of south Indian peninsular plains. It is well reputed in traditional system of medicine to

treat various ailments such as fevers, inflammations, skin infections and urinary infectio ns.

(Jeyachandran, Xavier, & Anand, 2003). In the Philippines, it is commonly known as

Makabuhay plant and can be seen in rainforests, nearby thickets and in most or all islands of

the country. There are tons of benefits of using the plant especially its stem as a primary

ingredient. The plant is also used in the Philippines for treating rheumatism and arthritis,

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 abdominal pains, scabies, skin ulcer and other diseases/ailments by mixing its extract with

coconut oil as traditionally done back then.

Staphylococcus aureus is a major bacterial human pathogen that causes a wide variety

of clinical manifestations. 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. S. aureus does not normally cause infection on healthy skin; however, if it

is allowed to enter the bloodstream or internal tissues, these bacteria may cause a variety of

potentially serious infections. Transmission is typically from direct contact. However, some

infections involve other transmission methods (Taylor & Unakal, 2017). It is the leading cause

of skin and soft tissue infections such as abscesses (boils), furuncles, and cellulitis. S. aureus can

cause serious infections such as bloodstream infections, pneumonia, or bone and joint

infections.

Considering the uses of T. cordifolia in traditional system of medicine, the researchers

will study on the antibacterial activity of T. cordifolia against Staphylococcus aureus in order

to know if the extract of the plant can inhibit the growth of this bacteria.

1.2 Theoretical Framework

Germs are present and can be found everywhere, it’s in the air, water and even the human

body isn’t spared of having it. There are major types of germs such as bacteria and fungi. Germ

Theory explains that some diseases are caused by the presence, actions and growth of

microorganisms within the body. It is considered as the cornerstone of modern medicine and

clinical microbiology. Germ theory led in to the development of antibiotics and hygie nic

practices. A lot of scientist and researchers played a huge part and contributed to this theory.

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 Louis Pasteur is one of the most major contributor of this theory. Pasteur is one of the first to

link germs to diseases. He also developed ways of making bacteria and viruses less dangerous.

It is relevant in this study to develop ways on how to treat, lessen and prevent the growth

of disease causing bacteria or organism in one’s body. With the help of Germ Theory, the

researchers are expected to prove that by the use of the extract of T. cordifolia, some plants have

antibacterial properties that can prevent and stop the growth of Staphylococcus aureus bacteria

in one’s body.

Figure 1.2 Theorertical Framework

1.3 Statement of the Problem

This research attempts to understand the antibacterial activity of T. cordifolia against

Staphylococcus aureus. More specifically it seeks to find the answers to the following

questions:

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 1. What are the antibacterial properties of Tinospora cordifolia?

2. Does the ethanolic extract of Tinospora cordifolia stem in 50%, 75%, and 100%

concentrations against Staphylococcus aureus capable of producing zone of inhibitio n

using Kirby-Bauer method?

3. Is there a significant difference between the 50%, 75%, and 100% concentratio ns

compared to Clindamycin in inhibiting Staphylococcus aureus?

1.4 Research Hypotheses

Based on the following questions, the following hypotheses are formulated:

1. Tinospora cordifolia contains antibacterial properties which are alkaloids,

glycosides, tannins, amino acids, carbohydrates, phenols and flavonoids.

2. The ethanolic extract of Tinospora cordifolia stem in 100%, 75% and 50%

concentrations are capable of inhibiting the growth of Staphylococcus aureus.

3. There is a significant difference between the 100%, 75% and 50% concentrations

compared to Clindamycin in inhibiting Staphylococcus aureus.

1.5 Significance of the Study

The findings in which this study will reveal, may benefit certain group and the benefits

they may be able to gain are as follows:

 Household

This study is beneficial to every household, especially to those who has T. cordifolia in

their backyard and is currently using it in many ways and purposes. It is hoped that this study

will encourage families to use a natural antibacterial agent in killing or stopping the growth of

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 harmful and dangerous micro-organisms, which can lead to serious major complications in the

near future.

 Consumers

Staphylococcus aureus is present in contaminated objects and is often spread through

skin-to-skin contact. This study aims to give knowledge or information if the consumers can

really use T. cordifolia in curing staph infections.

 Academe

This research aims to create a good image for the benefit of this institution. If the

findings and procedures stated were accurate and relevant, the school will gain credits.

 Researchers

This experimental research is a good opportunity to expand the knowledge of the

researchers that can help them in everyday living. The learnings obtain by the researchers can

also be used in conducting a new experimental research in the future and in using another plant

extract against a chosen bacteria.

 Future Researchers

The result of this research will serves as a good source of accurate and useful

information. In conducting new research or in testing the validity of other findings, the ideas

presented can be used as a reference. This study will also give them an overview about the

antibacterial property of Tinospora cordofilia, Kirby-Bauer method and Staphylococcus aureus.

1.6 Scope and Delimitations

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 This study will be conducted primarily to assess the antibacterial activity of Tinospora

cordifolia against Staphylococcus aureus. This will be tested using the Kirby–Bauer test where

according to Hudzicki (2009), the purpose of this is to measure the antibiotic sensitivity of

bacteria on an agar plate that is spread with bacteria and uses paper disks of antibiotics to test

the extent to which bacteria are affected by those antibiotics. Through experimental –

comparative method, the researchers can manipulate the amount of T. cordifolia stem extract

and ethanol. They are mixed to obtain the amount that has best to counter microorganisms. This

study is only limited to the antibacterial activity of ethanol extracts from the stems of T.

cordifolia to the bacteria Staphylococcus aureus. The study was conducted inside the Adamson

University Laboratory and shall be incubated for 24 hours to compare the diameter of inhibitio n

which forms around the paper discs where the best amount of antiseptic will show the largest

inhibition zone.

1.7 Definition of Terms

In order to have a better understanding for the readers and for clarification, the

researchers defined the important terminologies that will be seen in this study.

Antibacterial- destroying or inhibiting the growth of bacteria and especially pathogenic

microorganisms

Ethanol- a colorless volatile flammable liquid that has bactericidal activity and is used often as

a topical disinfectant. It is widely used as a solvent and preservative in pharmaceutica l

preparations as well as serving as the primary ingredient in alcoholic beverages

Infection- the act or result of affecting injuriously

Microorganism- an organism of microscopic or ultramicroscopic size

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Mueller Hinton Agar- Mueller and Hinton developed Mueller Hinton Agar (MHA) in 1941

for the isolation of pathogenic Neisseria species. Nowadays, it is more commonly used for the

routine susceptibility testing of non-fastidious microorganism by the Kirby-Bauer disk diffus io n

technique

Phytochemical Testing- refers to the extraction, screening and identification of the medicina lly

active substances found in plants. Some of the bioactive substances that can be derived from

plants are flavonoids, alkaloids, carotenoids, tannin, antioxidants and phenolic compounds

Staphylococcus- any of a genus (Staphylococcus) of non-motile gram-positive spherical

bacteria that occur singly, in pairs or tetrads, or in irregular clusters and include causative agents

of various diseases (such as skin infections, food poisoning, and endocarditis)

Tinospora cordifolia or Makabuhay plant- A plant that has a lot of medicinal claims such as

being a disinfectant. This is the main ingredient used as antiseptic agent to kill and prevent

growth of bacteria.

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 Chapter 2

REVIEW OF RELATED LITERATURE AND STUDIES

This chapter aims to find related studies and literature that will help the researches in conducting

their study. The researchers had found the following literature and studies as pertinent to the study

being discussed.

2.1 Biological and Phytochemical Analysis of Tinospora cordifolia

Stuart Jr. (2018) stated that the Tinospora codifolia has a wide range of use in terms of

its ability as an alternative medicine in which the stems and leaves of the plant are the main

parts that are utilized. Its properties are considered febrifuge, vulnerary, tonic, antimalaria l,

parasiticide, and insecticidal. Studies also suggests T. cordifolia is effective as an antioxida nt,

antidiarrheal, antimicrobial, anti-diabetic, anti-proliferative, cardiotonic, hypolipidemic,

antimalarial, ant nociceptive. Even though it has many uses, there are cautions in using the plant

especially those who are pregnant or patients with cardiac disorders.

According to Kaur, Lal and Suttee (2016), Tinospora cordifolia is one of the well-known

plants used for its medicinal use. In the present Pharmacological and Phytochemica l

investigation of Tinospora cordifolia it was standardized and compared as per World Health

Organization (WHO) guidelines. Different solvent extraction was done and then it was tested

for its active components. Preliminary phytochemical screening of Tinospora cordifolia showed

the presence of carbohydrates, glycosides, flavonoids, phenols, tannins and amino acids in the

crude drug.

The stem extracts exhibited dependent antimicrobial activity against both gram positive

and gram negative bacteria and can be used as a good therapeutic approach for infectious disease

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management and therapy. Tinospora cordifolia stem has shown different types of

phytochemicals and methanolic extract which exhibited a better antioxidant potential was found

more potent against both types of bacteria. Further studies on the Phyto-constituents and

evaluation of pharmacological activities of bioactive constituents of the drug was recommended

for future works.

According to Kaur, Prabhakar, Lal and Suttee (2016), the T. cordifolia stem has shown

different types of phytochemicals. T. cordifolia extracts showed the presence of carbohydrates,

glycosides, flavonoids, phenols, tannins and amino acids in the crude drug in the prelimina r y

phytochemical screening. The extracts of the stem of T. cordifolia showed the marked dose

dependent antimicrobial activity in vitro against both gram positive and gram negative bacteria

and it can be also used as a good therapeutic approach for infectious disease management and

therapy. The methanolic extract of T. cordifolia was found to be more potent against both the

group of bacteria, and also a better antioxidant.

According to Jyoti Rani, Lalita Singh, Hosihar Singh, Manish Kapoor and Gagandeep

Singh (2015), T. cordifolia leaf and stem extracts indicates the presence of alkaloids, cardiac

glycosides, tannins, phenols, carbohydrates and flavonoids. Jyotu Rani et al., suggested that

phytochemical component of T. cordolia is an importance source of bioactive compound that

can supply novel medicines and the phytochemical properties of T. cordifolia can be useful in

developing new specialized drugs.

According to Meenu (2016), active ingredients present in the T. cordifolia stem part

includes alkaloids, glycosides, steroids, and sesquiterpenoid. The pharmalogical activities of

the stem shows anti-osteoporotic activity, anti-inflammatory and wound healing activity, anti-

tuberculosis activity, and hepatoprotective and anti-oxidant activity. Thus, confirms the

9
therapeutic value of T. cordifolia. It also indicates that the chemical compounds of the plant

could serve as "lead" for developing agents for disorders in the coming years.

Patel & Ishnava (2014) cultured vegetative parts such as stem, leaf and nodal explants

from an elite in vivo grown mature plant of T. cordifolia on MS medium supplemented with

different hormonal concentrations and reported callus induction and organogenesis. Callus

formation occurred from nodal segments, leaf and inter-node explants when planted on differe nt

combinations of hormones. T. cordifolia showed response for in vitro shoot growth from the

nodal segment. The use of T. cordifolia for the treatment of general weakness and fever, the

immunomodulatory properties, hypoglycemic activities have been reported. Three major

groups of compounds— protoberberine alkaloids, terpenoids, and polysaccharides—ar e

considered as putative active constituents of T. cordifolia. Protoberberine alkaloids such as

berberine and palmatine are reported to have anti-cancer, anti-infective, anti-diabetic and

immunomodulatory activities.

According to Sinha, Mishra, Singh and Khanuja (2004), T. cordifolia is an important

drug in the Indian Medical System (IMS) and is used in medicines immemorial. The roots, stem

and leaves of the plant are also medicinal. In early times it was used as an Ayurvedic medicine

and is reported to be potent vegetable tonic. It was also used in folk and tribal medicine. It has

been established through modern testing and evaluation (pre-clinical and clinical trials) in

different disease conditions. These studies place this indigenous drug a novel candidate for bio

prospection and drug development for the treatment of such diseases as cancer, liver disorders,

ulcers, diabetes, heart diseases and postmenopausal syndrome where satisfactory cure

managements are still not available.

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 According to Choudhary (2018), T. cordifolia is a large, deciduous extensive ly

spreading climbing shrub with several elongated twining branches that is commonly found in

India. They grow in deciduous and dry forests. It has many medicinal properties like antibiotic,

immunosuppressant, anti-cancer, anti-spasmodic, anti-microbial, anti-osteoporotic, anti-

inflammatory, anti-arthritic, anti-allergic, anti-diabetic, anti-oxidant, anti-stress etc. A variety

of active components like alkaloids, steroids, diterpenoid lactones, aliphatics, and glycoside s,

have been isolated from the different parts of the plant body, including the root, stem, and leaves

of the plant. Guduchi is a traditional Ayurvedic medicine, and used in the treatment of fever,

diabetes, diarrhea, cancer, pain, skin disease etc.

According to Hook and Thomas (2018), T. cordifolia is a herbal medicine that has a lot

of use. It is a well-known medicinal plant and used to cure a number of disease. In today’s time,

it is also called as magical herb due to its property of curing a lot of diseases. It is commonly

used to cure migraine, asthma, fever, jaundice, diabetes, intestinal worms and piles.

According to Duraipandiyan, Ignacimuthu and Balakrishna (2012) T. cordifolia (Willd )

Miers (Guduchi) belonging to Menispermaceae (Family), which is native to and widely

distributed in the plains of India. Its plant, including the powder of its leaves, stem bark and

roots are used to treat ailments such as fever, diarrhea, cough, skin disease and a lot more in

folk and tribal medicine. Other than that it is used traditionally for treatment of inflammatio ns,

wound healing, antiseptics and fungal infections alike. With its diverse substance and natural

properties, T. cordifolia definitely has numerous benefits.

In a review on chemical and biological properties of T. cordifolia by Sankhala (2010) a

variety of chemical constituents isolated from the plant belong to different classes such as

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alkaloids, glycosides, diterpenoid, lactones, sesquiterpenoids and steroids that contains about

11.2% of protein and is rich in Ca and phosphorus. Along with that, it also contains some

biological properties that produces an antioxidant action mainly from its alcoholic root extract

where it is reported to have a defense mechanism in alloxan induced diabetic rats. It also

contains an Immuno- modulatory activity where both alcoholic and aqueous extract of

T.cordifolia have been tested successfully to improve the immune system and the body

resistance against infections. With the presence of ethanol, methanol and aqueous in the extract,

it has a wide range of antibacterial pathogens.

As stated by Saha & Ghosh S, (2012) T. cordifolia is a versatile resource for all forms

of life. The traditional ayurdendic medicine used it for ages in treating fever, jaundice, chronic

diarrhea, cancer, dysentery, bone fracture, pain, asthma, skin disease, poisonous insect, snake

bite, eye disorders. The report shows the active components of the plant such as alkaloids,

glycosides, lactones and steroids. These compounds have different biological roles of differe nt

types in targeting diseases. The major biological property of T. cordifolia include s

Immunomodulatory property, anti-diabetes property, anti-toxic effects, Anti-arthritic, anti-

osteoporotic effects etc. It also indicates that the site of action can lead us into identification of

novel perspectives into our understanding of nature.

In people’s everyday task in life, hands can always be covered with a lot of germs. It

may look clean in the outside but in reality it can cause illness and it can also spread to other

person by simply holding their hands even though it is washed. Many prevalent species of

bacteria can be found in our hands at any given time. The study of T. cordifolia: One Plant,

many roles by Saha and Ghosh (2012) showed the importance of the effectiveness of plant

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genetic resources that can help to prevent or to lessen the bacteria that can be the cause of illnes s

in our body.

According to a phytopharmacological review by Joshi and Kaur (2017) plants has been

one of the important sources of medicines since the beginning of human civilization. T.

cordifolia is a glabrous, succulent, woody climbing shrub native to India. It has anti- cancer,

nerve cell protecting, it is used to improve the immune system, the body resistance against

infections and most importantly it has a potential in preventing different diseases. The stem of

the plants is useful for skin problems and it is also used for treating wounds, pneumonia, asthma

and coughs.

Tinospora cordifolia, Nutritional,Ethnomedical and Therapeutic Utility by Kavya

(2015) stated that T. cordifolia belongs to a family Menispermaceae a well-known in Ayurveda

as Gudchi. It is a traditional plant medicine that contains an impressive level which in every

aspect it leads to a healthy and long life. There are many herbal plants in the world but Guduchi

has a higher medical value. Because of its stem and leaves it is used for treating various

ailments.

For the study of Sharma and Pandey (2010) entitled “Beneficial Effects of T.

cordifolia on Blood Profiles in Male Mice Exposed to Lead”, the stem and the leaves of T.

cordifolia does not just apply or treats humans but also animals. But it is widely used for humans

because of the bacteria and the diseases that most people encounter.

According to Mishra, Kumar and Pandey (2013), T. cordifolia has its medicinal efficac y

which includes phytochemical screening, antimicrobial, antioxidant, and anticancer activitie s

of the plant. Secondary metabolites including anthraquinones, terpenoids, and saponins were

13
 present in many extracts in addition to phenolics. Total phenol contents in various extracts were

found in the range of 8.75–52.50 catechol equivalent per gram. In disc diffusion assays, polar

extracts exhibited considerable inhibition against Klebsiella pneumoniae. Several other extracts

also showed antibacterial activity against pathogenic strains of E. coli, Pseudomonas spp.,

and Proteus spp. The study demonstrated the presence of various groups of phytochemica ls

in T. cordifolia extracts which are responsible for showing considerable antibacteria l,

antioxidant, and anticancer activities.

2.2 Antibacterial Activity of Tinospora cordifolia

Mohammed, Manish, and Dinesh (2012) stated that T. cordifolia occupies a very

important place in the field of medicinal plants and is widely used as a traditional medicine. It

is attributed to its ethno-medicinal uses since ancient times by traditional healers and physicia ns.

Local people use this plant for treating skin diseases, cleansing agent and many more. Since it

was also used as an antimicrobial plant, Mohammed, Manish, and Dinesh conducted a study on

the microbial property of T. cordifolia (roots). Based on their study even though the plant’s

ethanol and chloroform extracts have provided the best results as an antimicrobial this only

showed the basic information on the antimicrobial potential of the plant so further studies are

required to substantiate new biologically potent active antimicrobial compounds.

Razo et al (2015) stated that Makabuhay (T. cordifolia) plant has been identified to be

having antibacterial properties, giving rise to the formulated products such as ointments and

soaps were which are now commercially available. Since Staphylococcus aureus is the

causative agent of pimple and acne. The extract of T. cordifolia was experimented in differe nt

concentration against Staphylococcus aureus. The experiment confirms the antibacter ia l

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property of Makabuhay (T. cordifolia) plant extract as a formulated facial cleanser against

Staphylococcus aureus at different concentrations.

Aminul, Ashraful, and Mohammad (2011) has stated in their study the antimicrobia l,

cytotoxicity, and antioxidant activity of T. cordifolia. The stem was extracted with methano l

then partitioned before undergoing different experimentation. The results then indicated that the

extracts that were collected possess antioxidant, cytotoxic, and few antimicrobial substances

which justify the traditional uses of this plant in the treatment of different diseases. Furthermore

more future researches are necessary for elucidating the active principles.

According to the study of Singh, Panghal, Kadyan, Chaudhary, & Yadav (2014), that

Silver nanoparticles synthesized from T. cordifolia possess very good antibacterial activity

which makes them a potent source of antibacterial agent. The antibacterial activity of silver

nanoparticles prepared from T. cordifolia against multidrug resistant strains was determined by

agar well diffusion assay and Minimum Inhibitory Concentration (MIC) was estimated by

qualitative experimentation by resazurin based micro broth dilution method. The silver

nanoparticles of stem of T. cordifolia showed the zone of inhibition ranges from 10 ± 0.58 to

21 ± 0.25mm. The MIC of AgNPs from stem extract was found to be 6.25 to 200 μg/ml against

Pseudomonas aeruginosa (a courge of hospital burn units and its emergence as multidr ug-

resistant strains is a major problem in the control of nosocomial infection) strains.

The antibacterial activity of the aqueous, ethanol and chloroform extracts from the stems

of T. cordifolia was studied using disc diffusion method against Escherichia coli, Proteus

vulgaris, Enterobacter faecalis, Salmonella typhi (Gram-negative), Staphylococcus aureus and

Serratia marcesenses (Gram-positive). Previous study of Jeyachandran, Xavier, & Anand

15
(2003) suggest that the ethanolic extract has significant antibacterial activity against tested

bacteria. The study justifies the claimed uses of T. cordifolia in the traditional system of

medicine to treat various infectious diseases.

The medicinal plants are used in traditional cure of various diseases and an impressive

number of modern drugs have been isolated from these natural resources, many based on their

use in traditional medicine. Plants used in traditional medicines, contain a vast array of

substances that can be used to treat chronic and infectious diseases. The antibacterial activitie s

of extracts of stem and leaves of T. cordifolia were investigated to evaluate its medicina l

potential. The extracts were screened against Escherichia coli, Bacillus subtilis, Pseudomonas

aeruginosa and Staphylococcus aureus by disc diffusion method. Based on the study

conducted by Verma & Rojan (2012) the methanol extracts of T. cordifolia was active on all

the strains of pathogens. The result of the study validates the use of methanol extract of this

species in ethnomedicine and could provide a lead in the isolation of antibacterial agents from

methanol extract of T. cordifolia.

According to Singh, Katiyar, Singh and Srivastava (2015), T. cordifolia can be a good

and potential source for herbal drug preparations in treating UTI. The antimicrobial activitie s

of T. cordifolia extracts against the pathogens causing urinary tract infections were examined

in the study. T. cordifolia is one of the most versatile herbs. It is also called as nectar of life, as

it is extremely useful in curing and strengthening the immune system of the body. The extract

of the plant contains several bitter components such as glucosides, alkaloids, a glycoside-gilo in,

a non-glucoside-gilenin, gilosterol, alkakoid tinosporin, tinosporic acid, tinosporol, berberine,

tinosporidine, sitosterol isolated, cordifol, heptacosanol, octacosonal and a new furano id

16
diterpene-tinosporide. The stem of T. cordifolia is used as an ingredient in Ayurvedic

preparations used in general debility, dyspepsia, fevers and urinary diseases. The use of plant

extracts with known antimicrobial properties, can be of great significance in therapeutic

treatments.

A study conducted by Hipol, and Cariaga (2012) entitled “Anti-inflammatory Activitie s

of the Aqueous Extract of the Stem of Tinospora crispa (Family Menispermaceae)” aims to

evaluate the anti-inflammatory properties of the extract of stem of Tinospora crispa. Forty five

albino rats (Rattus norvegicus) of either sex weighing 180-200g were used for the Carageenan-

induced paw edema assay. Tinospora crispa was found to cause stabilization of cellula r

membranes at 5%w/v and 7.5% w/v concentrations and inhibition of protein denaturatio n.

These activities are therefore suggested as the possible mechanisms for the anti-inflammato r y

action of Tinospora crispa. The experiment on the reduction of induced rat paw edema evidently

confirmed the anti-inflammatory activity of Tinospora crispa extracts.

Mutalik M. & Mutalik N. (2011) showed the links between what is believed and what

is known regarding the use of T. cordifolia. Evidences from various pharmacological studies

suggest beneficial effects. Immunomodulation, protective actions, and antioxidant properties

helps in treating diabetes, obstructive jaundice, malaria, hepatic and splenic injury, protection

from allergens and toxins, infections, inflammation, rheumatoid arthritis, gout, leprosy, and in

preventing cancer and tuberculosis.

Castillo, et al. (2013) evaluated the clinical efficacy and safety of T. cordifolia lotion

including its cure rate and clearance time compared with permethrin lotion. A clinical study was

performed in three government institutions to investigate clinical efficacy of T. cordifolia lotion

17
in sixty-six scabies-infected patients. The patients were treated with T. cordifolia or permethr in

lotions for three consecutive days for two weeks and clinical assessment of each patient was

performed for five weeks. The researchers found out that T. cordifolia lotion exhibits anti-

scabies activity comparable with permethrin.

Arcueno, Retumban, Echano & Guerrero (2015) aimed to compare the effect of

Tinospora cordifolia on healing of diabetic wound among albino mice when given and

administered along with an ointment topically applied to the wound. Results showed a

significant reduction in blood glucose level among mice given the plant extract.

To conclude, Patil, Kulkarni and Pandey (2017) stated that the T. cordifolia extract have

been reported to have potential against microbial infections. This was tested against gram

positive and gram negative organisms such as E. coli and S. Aureus with the use of disc diffus io n

method and gained results that shows antifungal activity among the entire fungal organism.

Kumar et al. (2017) in their study revealed that T. cordifolia leaves were tested and

observed on Escherichia coli on three different solvent. Ethanolic, methanolic and aqueous

extracts of the leaf were tested using slip disc method to amount the anti-microbial against E.

coli. As the result, the medicinal plant proves that it can be an alternative form of medicine than

the consumers used with its fewer side effects and easy availability.

According to Nagaprashanthi et al. (2012), the present investigation was carried out to

evaluate the in vitro antifungal and antibacterial activity of hydro alcoholic extract of T.

cordifolia creeped on Azadirachta indica Tree (TC1) in comparison with that of T. cordifolia

(TC2) creeped on fencing. Hydroalcoholic extract of T.cordifolia stem was prepared by

maceration technique. The microorganisms used as antibacterial and antifungal were

18
Staphylococcus aureus, Escherichia coli, Klebsiella pneumonia, Pseudomonas sp, Aspergillus

niger, Aspergillus fumigates, mucor sp and Pencillium. The extract of TC1 exhibit effective

antimicrobial activity against all the organisms, while the extract of TC2 exhibits inhibitio n

zone on limited species such like Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas

sp, Aspergillus niger, Aspergillus fumigates, and mucor sp. The researchers concluded from

this study that T. cordifolia stem extract creping on Azadirachta indica has antimicrob ia l

activity similar that is similar to neem tree when compared to T. cordifolia creping on fencing.

This can explain that the host plant will gain some of the activities when they survive on

medicinal plants.

According to Patil, Kulkarni and Pandey (2017), the present work was being carried out

to isolate and identify the active constituents of the plants responsible for antibacterial activity.

The aqueous, methanol, ethanol and acetone extract of T. cordifolia, Azarchita indica and

Ocimum santum leaves extract were screened for the presence of phytochemical components

and tested for antibacterial activity against Escherichia coli, Staphylococcus aureus, Klebsiella

pneumonia, Enterobacter aerogenes, Pseudomonas aeruginosa, Salmonella typhimurium,

Salmonella typhi, Staphylococcus epidermidis and Proteus vulgaris. The results obtained in this

study thus suggests that the identified phytochemicals may be the bioactive constitue nts

responsible for the efficacy of leaves extract of T. cordifolia, Azarchita indica and Ocimum

santum against fever, syphilitic, ulcer, inflammatory disease wounds, conjunctivitis etc. It

suggested that the traditional medicinal use of T. cordifolia, Azarchita indica and Ocimum

santum be continued and have a scientific evaluation of its active constituents given serious

attention.

19
 This study aims to investigate the in vitro antibacterial activity of T. cordifolia stem

extract against Escherichia coli O78. According to Mamta and Jakhar (2016), medicinal plants

have been clinically used in curing various human and animal disorders. T. cordifolia is one of

the most commercially exploited plants in pharmaceuticals. The estimated annual demand of

this species used in the preparation of crude herbal drugs in the Indian system of medicine s

(ISM) is 10,000 tonnes. The stem pieces were washed, shade dried and powdered in a mixer.

For preparation of extract, 15 g of powdered material was boiled in 100 ml of water and filter ed

through Whatman`s No.1 filter paper (Singh et al., 2014). The extract was used for assessment

of in vitro antibacterial activity of T. cordifolia stem extract against Escherichia coli O78. The

overall results of in vitro antibacterial activity of aqueous extracts of T. cordifolia against E.

coli indicated that the extract has potential to be used commercially for control of E. coli

infection.

2.2 Staphylococcus aureus

The Staphylococcus aureus bacterium, commonly known as staph, was discovered in

the 1880s. In the 1940s, medical treatment for S. aureus infections became routine and

successful with the discovery and introduction of antibiotic medicine, such as penicillin.

According to Stoppler (2018), the name Staphylococcus comes from the Greek staphyle,

meaning a bunch of grapes, and kokkos, meaning berry, and that is what staph bacteria look like

under the microscope, like a bunch of grapes or little round berries. It is considered as one of

the major human pathogen and the most common community-acquired etiological agents and

nosocomial bacterial infection. Other than that, it is an important opportunistic pathogen, where

it is rarely affecting those patients whose immune system is depressed, responsible for a variety

20
of diseases ranging from minor skin infections to life-threatening systemic infections. It is one

of the common bacterial infections in humans.

Staphylococcus is a group of bacteria that can cause a multitude of diseases and can

cause a number of infectious diseases in various tissues of the body. Staph infections are caused

by the bacteria staphylococcus that is most of the time, cause no problems or result to minor

skin contamination. But, these infections can in turn be deadly if the bacteria invade deeper into

the human body such as bloodstream, heart and lungs. According to Taylor and Unaka (2017)

these bacteria are present in up to 25% of healthy people and can be found in the normal human

flora associated with the skin and mucous membranes of every human being. Over 30 differe nt

types of staphylococci can infect humans, but most infections are caused by Staphylococcus

aureus. Common symptoms such as Boils under the arms or around the groin or buttocks, a

painful rash caused by Impetigo, Cellulitis and the Staphylococcal scalded skin syndrome which

affects mostly babies and children that features fever, rash and blisters. Food poisoning may

also be caused by these bacteria where its symptoms come and disappear quickly, often lasting

for half a day. It also causes Septicemia and Bacteremia, known as blood poisoning where the

bacteria enter a person’s bloodstream. Therefore, Staph contaminations are infectious until the

point when the disease has settled. Direct contact with an infected sore, wound, or with personal

care items are common routes of transmission.

In the study of Espedido, et al. (2012) Staphylococcus aureus bacteremia is an infectio n

with an incident rate ranging from 20% to 50% cases per year. Patients die from SAB between

10% and 30%. Pathogen-host interactions, especially the presence of shock and the source of

SAB, are strong predictors of outcomes. At this rate, this account for a greater number of deaths

than AIDS, and tuberculosis. Although antibiotic resistance was associated, pathogen-specific

21
 factors and poor response to antibiotic therapy was observed. The attributable mortality of S.

aureus bacteremia (SAB) is as at yet higher and indicators for mortality and clinical results of

this condition are in need to be clarified. In patients with Staphylococcus aureus bacteremia,

infectious disease consultation remains a valuable device to enhance clinical result.

Harmful microorganisms can be transferred to hands from contaminated surfaces

individuals come into contact in day by day life. These kind of surfaces contribute a lot to the

bacterial load of hands which in turn transmit illness to one self and in addition to others. A

study by De Alwis et al. (2012) showed that with 60 selected medical students taken randoml y

for bacterial count from both hands before and after toilet use, bacterial load in the hands for

both males and females showed an increment after toilet use. The bacteria Staphylococcus

aureus mainly appeared to isolate the hands of selected students the most.

2.4 Synthesis

Tinospora cordifolia is a large, deciduous extensively spreading climbing shrub with

several elongated twinning branches that is commonly found in India (Choudhary 2018). All

the parts of this plant has its own use and purpose starting from its leaves down to the roots.

Tinospora cordifolia has a lot of medicinal claims (Mishra, Kumar and Pondey 2013). Before

it was used to treat ailments such as fever, diarrhea and cough and used as tribal medicine. Now

in today’s time, its stem is usually used for treating pneumonia and asthma (Joshi and Kaur

2017). Stem extract of Tinospora cordifolia has active components and shows presence of

carbohydrates, glycosides, flavonoids, phenols, tannins and amino acids in the crude drug in its

preliminary phytochemical screening (Kaur, Lal and Suttee 2016). Alkaloids, steroids, and

sesquiterpenoid are also active in its stem that can serve as a lead for developing medicines in

the future (Meenu Bhan's Phytochemical and pharmacological evaluation 2016). Some of its

22
 common medicinal claims is that it’s used as an antioxidant, anticancer and most especially as

an antimicrobial (Mishra, Kumar and Pondey 2013). Because of being identified as having

antibacterial property, Tinospora cordifolia is now giving rise to formulated products such as

ointments and soaps which are now commercially available (Razo 2015). Tinospora cordifolia

was tested to identify the active constituents of the plants antibacterial activity against gram

positive and negative organisms such as E. coli and Staphylococus aureus (Patil, Kulkarni and

Pandey 2017). Staphylococus aureus is one of the common bacterial infections in humans

(Stoppler 2018). It can cause multiple number of infectious disease in the body. It can be

transferred to hands from contaminated surfaces of individuals in our daily life. With Tinospora

cordifolia, we can now prevent harmful disease causing bacteria like Staphylococus aureus

from spreading.

2.5 Conceptual Framework

This experimental study focuses on the antibacterial activity of Tinospora cordifolia

against Staphylococcus aureus. This section explains the designed framework of the whole

dissertation and explain the relationship among different concepts in the study.

Figure 1 shows the conceptual framework of the study. The input is the title of the

study. It involves the variables of the study.

Then the process which includes the materials for the disk-diffusion method. The

researchers will culture the Staphylococcus aureus bacteria using blood agar.

23
 The output includes the experimentation itself. The researchers will used the Disk-

diffusion method and measure the zone of inhibition. The statistical analysis specifically,

Analysis of Variance (ANOVA), will be used to determine the differences of the samples.

Figure 2.5 Conceptual Framework

24
 Chapter 3

RESEARCH METHODOLOGY

This chapter contains the research design, locale, materials and techniques that will be used to

be able to come up with a result.

3.1 Research Design

The research study is experimental in nature that aims to determine the antibacter ia l

activity of T. cordifolia against Staphylococcus aureus. The specific type of bacteria to use is

the Staphylococcus aureus. A phytochemical testing was done to determine the active

components present in T. cordifolia. The concentrations of T. cordifolia extract are 100%, 75%,

and 50% that were produced from simple dilution. Kirby-Bauer method was used in this study,

to determine the resistance mechanism and susceptibility of Staphylococcus aureus against the

T. cordifolia compared to Clindamycin which is the positive control and distilled water which

is the negative control. Kirby-Bauer testing measures sensitivity of bacteria to antibiotics by

culturing bacteria on solid growth media surrounding sources of drug. The zone of inhibition is

an area of clear media where bacteria are not able to grow surrounds the wafer. A lager zone of

inhibition around an antibiotic-containing disk indicates that the bacteria are more sensitive to

the antibiotic in the disk.

3.2 Material and Study Site

The main materials that will be used are the ethanolic extract of T. cordifolia in 100%,

75% and 50% concentrations, Staphylococcus aureus bacterium, one capsule of Clindamyc in

and distilled water. The study was held in Adamson University located at San Marcelino St,

25
 Ermita, Manila, Metro Manila during the first semester of the school year 2018-2019. The

extraction and the phytochemical testing of T. cordifolia plant was also held at Adamson

University Biology Laboratory.

3.3 Data Collecting Instruments

The following instruments that will be used in this study are specified in this section.

Alcohol Lamp Petri Dish

Autoclave Puncher

Beaker Stirring rod

Biosafety Cabinet Level 2 Test tube

Digital caliper Test tube rack

Erlenmeyer Flask Whatman no. 1 and 2

Forceps Heat Plate

Inoculating Loop Incubator

Table 3.3 Instruments

Alcohol lamp- Alcohol lamp will be used to sterilize the inoculating loop and specimen to avoid

contamination.

Autoclave- Autoclave will be used to sterilize laboratory instruments in order to eliminate

remnants of bacteria.

Beaker- Beaker will be used to measure the amount of distilled water needed for the preparation

of media.

Biosafety Cabinet Level 2- Biosafety Cabinet Level 2 was used as a working site for the

experimentation. Wherein mild and potential bacteria is being isolated to avoid being infected.

26
Digital caliper- Digital caliper will be used to accurately measure the distances of zones of

inhibition produced by the disks.

Erlenmeyer flask- Erlenmeyer flask will contain the solution for the media preparation. It will

then be placed in a heat plate for the solution to be dissolved.

Forceps- Forceps will be used to transfer 6mm disks to the petri dish.

Heat plate- Heat plate will be used to dissolve the solution for media preparation. It is important

that the solution will be dissolved properly to obtain the standard of the media and its efficacy.

Incubator- Incubator will be used to assist the growth of the culture. The specimen is usuall y

incubated at 37°C for 24 hours or depending on the growth requirement of the organism.

Inoculating loop- Inoculating loop will be used to inoculate the specimen into the media. It is

important to sterilize it first with alcohol lamp to avoid contaminatio n.

Petri dish- Glass and disposable plastic petri dish of standard size will contain the media of

choice.

Puncher- 6 mm diameter puncher was used to produce blank disks.

Stirring rod- Stirring rod aids to the dissolving of the solution for media preparation.

Test tubes- Test tubes will be used to fill and prepare the bacteria for sterile swabbing.

Test tube rack- Test tube rack will be used to hold and store test tubes that contains bacteria.

Whatman no. 1 and 2- The 6mm disks for Kirby-Bauer method was prepared using

Whatman filter paper no. 1 and 2.

27
3.4 Data Gathering Procedure

 Tinospora cordifolia Extraction and Preparation

The plant was collected at San Nicolas, Arayat, Pampanga Philippines. An adequate

amount of plant were brought to Adamson University to undergo a process of extraction and

phytochemical testing. The T. cordifolia stems were air dried then were cut into portions. The

stems were grinded using a blender and the dried powder that was collected was stored in an

airtight and light resistant bottle.

 Tinospora cordifolia Ethanolic Extract

The powdered T. cordifolia stems were brought to the Adamson University where it will

be incorporated with ethyl alcohol. The filtrate was evaporated using the rotary evaporator to

yield the ethanolic extract of T. cordifolia,

 Phytochemical Components of Tinospora cordifolia

The phytochemical analysis of T. cordifolia were also done at Adamson Univers ity

where it was tested to determine the active components which are alkaloids, flavonoid s,

glycosides, tannins and amino acids present in the plant.

Detection of Alkaloid

Extracts were dissolved individually in dilute Hydrochloric acid and filtered.

Hager’s Test: Filtrates were treated with Hager’s reagent (saturated picric acid solutio n).

Presence of alkaloids confirmed by the formation of yellow coloured precipitate.

Detection of Flavonoids

28
 Alkaline Reagent Test: Extracts were treated with few drops of sodium hydroxide solutio n.

Formation of intense yellow colour, which becomes colourless on addition of dilute acid,

indicates the presence of flavonoids.

Detection of Glycosides

Keller-Killani Test: Extract were treated with 2ml of glacial acetic acid containing one drop

of ferric chloride solution. Appearance of a violet ring below the brown ring and a greenish ring

in the acetic acid layer confirmed the results.

Detection of Tannins

Extract was boiled in 20 mL distilled water and few drops of ferric chloride were added to the

filtrate; a blue-black precipitate indicated the presence of Tannins.

Detection of Amino Acids

Millon’s Test: Filtrates were treated with few drops of Millon’s reagent. A white precipitate is

formed indicating the presence of amino acids.

 Tinospora cordifolia Concentrations

The T. cordifolia ethanolic extract will be further diluted with distilled water to achieve

the desired concentrations of 100%, 75% and 50%. For the ratio of each concentrations of the

extract, the 100% concentration contains purely 10ml of the extract of T. cordifolia. The 75%

concentration contains 15ml of the ethanolic extract of T. cordifolia and 5ml of distilled water

while the 50% concentration contains 10ml of ethanolic extract of T. cordifolia and 10ml of

distilled water.

 Staphylococcus aureus Preparation

A pure culture of Staphylococcus aureus will be coming from Adamson. Incubation of

the specimen will be done for 24 hours at 37 ° C or at room temperature.

29
 Preparation of Mueller-Hinton agar

Mueller-Hinton agar is commonly used in disk diffusion test or Kirby-Bauer test. Kirby-

Bauer method determines the resistance or sensitivity of facultative anaerobes which can be

used to determine the treatment for bacterial infections.

In standard preparation of Mueller-Hinton agar, 38.0 grams of the medium was added

to 1 liter of distilled water. Then, it was boiled for 1 minute with frequent agitation to complete ly

dissolve the medium. Following that procedure, it was autoclaved for 1 hour at 15 PSI (121°C)

and will then be cooled at room temperature (HiMedia Laboratories Pvt., Mueller-Hinton Broth,

2017).

 Kirby-Bauer method/ Disk Diffusion Method

Kirby-Bauer method, also known as the Disk Diffusion method, is used to determine

the ability of a specific antibiotic to inhibit a particular bacterium. The result of this method is

shown by the different zones of inhibition produced around the disks which may be reported as

susceptible (able to inhibit the bacteria), intermediate, or resistant (unable to inhibit the

bacteria).

The disks used in this study were prepared from a Whatman filter paper no. 1 and 2

punched into 6 mm disks. The disks were autoclaved for sterilization. After it was incubated,

the prepared extracts with predetermined concentrations were incorporated into the disks. This

study used Clindamycin as the positive control, and distilled water incorporated into blank disks

as the negative control.

30
 Figure 3.4 Methodology Flowchart

3.5 Data Analysis

The researchers will used statistical analysis specifically ANOVA (Analysis of

Variance). ANOVA will be used to determine whether the 100%, 75% and 50% concentratio ns

compared to clindamycin are significantly different with each other. The researchers also used

descriptive statistics to determine the mean and standard deviation.

3.6 Ethical Considerations

There will be no humans or animals that could be subjected to harm in any ways. S.

aureus is the only test subject that the researchers will focus on. Since S. aureus is pathogenic

to humans, the researchers will ensure that their qualifications are adequate for safe conduct of

the study. All of the instruments will be sterilized before and after the experiment. The

researchers acknowledge the works of other authors that were used in this research paper. The

presentation of data findings will not be biased and there would be no misleading information.

31
 Chapter 4

PRESENTATION, ANALYSIS, AND INTERPRETATION OF DATA

In this chapter, the collected data will be finally discussed and presented based on the

researchers’ analysis. Since the research design of this study is experimental, the researchers gathered

the data based on the experiment they have done. All of the data that are presented here are purely from

the experiment.

The data are classified into three major divisions or themes, based on the statement of the

problem as seen on the first chapter. First is the Phytochemical Analysis, then the Disk Diffus io n

Method and lastly the Data Analysis that the researchers used in the study which is ANOVA or Analysis

of Variance.

4.1 Antibacterial Properties of Tinospora cordifolia

Ethanolic extract of Tinospora cordifolia underwent various chemical tests to detect the

chemical constituents present in them.

ANALYSIS OF PHYTOCHEMICALS PRESENT IN THE ETHANOLIC EXTRACT OF

Tinospora cordifolia
(Notation: (+) = Present and (-) = Absent)
Secondary Metabolite Result

Flavonoids +

Alkaloids +

Tannins +

Glycosides +

Amino Acids +

Table 4.1: Phytochemical Analysis of the Ethanolic extract of Tinospora cordifolia

32
 Phytochemical investigation of the ethanolic extract of Tinospora cordifolia stems led

to the presence of flavonoids, alkaloids, tannins, glycosides and amino acids. Jyotu Rani et al.

(2016) concluded that the phytochemical properties of T. cordifolia is an important source of

bioactive compound that can supply novel medicines and can be useful in developing new

specialized drugs.

4.2 Zone of Inhibitions

Guideline Chart for the Susceptibility of Staphylococcus aureus to Tinospora cordifolia

Resistant Intermediate Susceptible

(< or = mm) (mm) (= or > mm)
14 15-20 21

Table 4.2.1: Guideline chart for susceptibility

Table above shows the guideline chart for the disk diffusion method. Zone of inhibitio ns

measuring less than or equal to 14mm indicates resistance of the bacteria. The sensitivity of

bacterial strains is said to be intermediate if the zone of inhibitions measures from 15mm-20mm.

Zone of inhibitions indicating susceptibility of the bacteria will measure equal or greater than

21.

Zone of Inhibition of the Distilled Water

Trial 1 Trial 2 Trial 3

Distilled Water 0.00 mm 0.00mm 0.0mm

Table 4.2.2: Zone of Inhibition of the negative control

33
 The disks incorporated with distilled water, which is the negative control were not able

to produce zone of inhibition, indicating the resistance of the bacteria. Three trials were

performed and results showed 0.00mm zone of inhibition.

Zone of Inhibition of Clindamycin

Trial 1 Trial 2 Trial 3

Clindamycin 31.79 mm 33.22 mm 32.88 mm

Table 4.2.3: Zone of Inhibition of the positive control

The disks incorporated with clindamycin which is the positive control showed

susceptibility of the bacteria. In the first trial the zone if inhibition is 31.79mm and in the second

trial the zone of inhibition is 33.22mm and the zone of inhibition for the third trial is 32.88mm.

Zone of inhibition of the 100%, 75% and 50% concentrations of the ethanolic extract of

Tinospora cordifolia

Trial 1 Trial 2 Trial 3

100% Ethanolic
23.38 mm
extract of Tinospora 17.21mm 26.81 mm

cordifolia
75% Ethanolic
0.00 mm 0.00 mm 0.00 mm
extract of Tinospora
cordifolia
50% Ethanolic
0.00 mm 0.00 mm 0.00 mm
extract of Tinospora
cordifolia
Table 4.2.4: Zone of inhibition of the different concentrations

34
 Data shown above shows the different zone of inhibition. Three trials were performed

for each control. The disk incorporated with 100% concentration of the ethanolic extract of

Tinospora cordifolia were able to produce measurable zone of inhibitions which are 23.38mm,

17.21mm and 26.81mm , indicating the susceptibility of the bacteria. The disk incorporated

with 75% concentration and 50% concentration of the ethanolic extract of Tinospora cordifolia

were not able to produce measurable zone of inhibitions, indicating the resistance of the

bacteria.

4.3 Analysis of Variance (ANOVA)

SUMMARY 100% 75% 50% Total

Ethanolic Extract of Tinospora cordifolia

Count 3 3 3 9
Sum 67.4 0 0 67.4
Average 22.46666667 0 0 7.488889
Variance 23.66563333 0 0 132.104186
Table 4.3.1: Summary of the count, sum, average and variance of the concentrations

This table summarizes the collected data from the ethanolic extract of T. cordifolia in

different concentrations. The three different concentrations have a total yield of 67.4mm, an

average of 7.5 and a variance of 132.

Distilled Water

Count 3 3 3 9
Sum 0 0 0 0
Average 0 0 0 0
Variance 0 0 0 0
Table 4.3.2: Summary of the count, sum, average and variance of the distilled water

This table summarizes the collected data from the negative control which is the distilled

water. The three trials have a total yield of 0.mm, an average of 0 and a variance of 0 for there

were no zone of inhibitions produced.

35
 Clindamycin

Count 3 3 3 9
Sum 97.89 0 0 97.89
Average 32.63 0 0 10.8766667
Variance 0.5581 0 0 266.31875
Table 4.3.3: Summary of the count, sum, average and variance of the clindamycin

This table summarizes the collected data from the positive control which is the

clindamycin. The three trials have a total yield of 97.89mm, an average of 10.88 and a variance

of 266.32.

Descriptive Statistics
Dependent Variable: Zone of inhibition

Control Mean Std. Deviation N

100% extract 22.4667 4.86473 3

50% extract .0000 .00000 3

75% extract .0000 .00000 3

Clindamycin 32.6300 .74706 3

Distilled water .0000 .00000 3

Total 11.0193 14.48020 15

Table 4.3.4: Descriptive Statistics

This table shows the mean of the different independent variable. The mean of the 100%

ethanolic extract of T. cordifolia is 22.47 while the mean of the distilled water, 75% and 50%

ethanolic extract of T. cordifolia is 0. The clindamycin has a mean of 32.63. The total mean is

11.02.

36
 Figure 4.3: Estimated Marginal Means

The graph shows the relationship between the different controls used in the study. The

clindamycin has the highest marginal mean which is greater than 30mm. The 100%

concentration of the ethanolic extract of T. cordifolia has an estimated marginal mean of greater

than 20. And lastly, for the distilled water and the 75% and 50% ethanolic extract of T. cordifolia

showed 0 marginal mean.

ANOVA
Source of
Variation SS df MS F P-value F crit
Sample 557.5871185 2 278.793559 103.581971 1.333E-10 3.554557
Columns 2023.761785 2 1011.88089 375.950639 2.087E-15 3.554557
Interaction 1115.174237 4 278.793559 103.581971 3.619E-12 2.927744
Within 48.44746667 18 2.69152593

Total 3744.970607 26
Table 4.3.4: Analysis of Variance

37
Alternative Hypothesis: There is a significant difference between the 50%, 75%, and 100%

concentrations compared to Clindamycin in inhibiting Staphylococcus aureus.

Null Hypothesis: There is no significant difference between the 50%, 75%, and 100%

concentrations compared to Clindamycin in inhibiting Staphylococcus aureus.

The P-value helps you to determine the significance of the results. A small p-value

(typically ≤0.05) indicated strong evidence against the null hypothesis, so you reject the null

hypothesis. According to the table above, the p-values are less than .05. Therefore, the

researchers reject the null hypothesis since there is a significant difference between the 100%

concentration of the ethanolic extract of Tinospora cordifolia and clindamycin to the distilled

water and the 75%, 50% concentrations of the ethanolic extract of Tinospora cordifolia.

38
 Chapter 5

SUMMARY OF FINDINGS, CONCLUSIONS, AND RECOMMENDATIONS

This chapter includes the summary of the findings that are collected, the formula ted

conclusions and the recommendations for the study.

5.1 Summary of Findings

This includes the summary of findings or result gathered through the duration of the study.

1. The ethanolic extract of Tinospora cordifolia underwent various chemical tests to detect

the chemical constituents present in them. Through this phytochemical test of the

extract, it was determined to have active constituents such as flavonoids, tannins,

glycosides, amino acids and alkaloids which are bioactive compounds/compone nts

found in plant. Glycosides, amino acids, tannins, flavonoids and alkaloids are active

constituents of Tinospora cordifolia for its antibacterial activity.

2. 100%, 75%, and 50% concentrations of the ethanolic extract were formulated by adding

distilled water. After using the Kirby- Bauer method, the 75% and 50% concentratio ns

weren’t able to show zone of inhibition while the extract with the 100% concentratio n

was observed as the one which successfully exhibit the antibacterial activity of

Tinospora cordifolia against the gram-positive bacteria S.aureus.

3. In the 100%, 75% and 50% concentrations of the ethanolic extract of Tinospora

cordifolia, the 75 % and 50% concentrations were not as successful in inhibiting

Staphylococcus aureus bacteria while the 100% concentration showed a significa nt

result of inhibition closer to the positive control which is Clindamycin.

39
 5.2 Conclusions

The following are the conclusions formulated from the findings in the experiment that

was conducted.

1. The active constituents present in Tinospora cordifolia were flavonoids, tannins, amino

acids, glycosides and alkaloids. These constituents are responsible for the antimicrobia l/

antibacterial activity of the plant.

2. The determination of antibacterial activity of Tinospora cordifolia extract

concentrations 100%, 75% and 50% with the use of Kirby-Bauer method only showed

zone of inhibition in the 100% concentration, an indication of susceptibility of the

bacteria against a specific concentration of Tinospora cordifolia.

3. In comparison with clindamycin, 75 % and 50% concentrations were not successful in

inhibiting S.aureus bacteria while the 100% concentration showed a significant result of

inhibition closer to the positive control which is Clindamycin.

5.3 Recommendations

The following are the recommendations that the researchers would like to have for the future

researchers that would like to use this study as their reference.

1. The researchers’ would like to recommend the use of other parts of the Tinospora

cordifolia plant in testing its antibacterial/antimicrobial activity aside from the stems

which were used in the study.

2. The researchers’ would like to recommend to the future researchers to test the

susceptibility of the plant against gram-negative bacteria or any bacteria aside from

S.aureus.

40
3. The researchers’ would like to recommend to the future researchers to have additiona l

phytochemical tests of active components of the plant aside from the ones done in the

study.

4. The researchers’ would like to recommend the use of Tinospora cordifolia as an

antibacterial or antimicrobial agent.

5. The researchers’ would like to recommend the further development of Tinospora

cordifolia and eventually turn it into commercially used products such as antibacter ia l

agents.

41
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Kumar,D.V, Geethanjali,B., Avinash,K., Kumar,J.R, & Chandrashekrappa,G. (2017).Tinospora
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Nagaprashanthi,C., Rafi khan.P, Gopi chand.K, Aleemuddin MA,and Rajiya Begum.G. (2012).
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Patil,P., Kulkarni,C., & Pandey,A. (2017).Antibacterial and phytochemical analysis of Tinos
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2249 – 4340

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Singh, K., Panghal, M., Kadyan, S., Chaudhary, U., & Yadav, J. (2014). Antibacterial Activity of
Synthesized Silver Nanoparticles from T. cordifolia against Multi Drug Resistant
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 isolated-burn-patients-2157-7439-5-192.php?aid=25859

Singh, P., Katiyar, D., Singh, B. & Srivastava, A. (2015). Antimicrobial activity of Tinospora
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UpadhyayA. K.,Kumar K., Kumar A.,& Mishra H.S. (2010). T. cordifolia (Willd.)Hook.
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cordifolia Miers. Extracts Against Dental Pathogens. Journal of Pharmacology and
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46
 Appendix A
PERMIT TO CONDUCT RESEARCH

Dr. Ramon P. Luber, DVM, MSc

Chairperson, Biology Department

Name of thesis: “The Antibacterial Activity of Tinospora cordifolia against Staphylococcus

aureus”
Contact information of each group member:
Name of Student Contact Number Email Address

Alcyren Joy De Guzman 09351418980 deguzman.alcyrenjoy@gmail.com

Alliah Lorio 09455611272 lorioalliahb3@gmail.com
Jasmine Nituda 09950801509 jasminenituda@gmail.com
Nyzel Baydo 09367200198 nyzeeelbyd@gmail.com
Giselle De Guzman 09178251114 dgisellejane0514@gmail.com

Faye Baricaua 09663411557 mayflor.baricaua611@yahoo.com

Mernelle Santos 09293377263 mernelle_mae@yahoo.com
Sofia Landrito 09475342531 sofialubelle17@gmail.com
Slinnie Enciso 09569710305 slinniechan@gmail.com

Noted and Approved by:

Mr. Glen S. Mirabete Mrs. Rebecca Martillana-Baloloy

Research Adviser/Teacher Academic Coordinator

47
 Appendix B
RRL SUMMARY

RRL Summary
August 13 2018
Date:
Month Day Year

BUILDING LITERATURE REVIEW

BARICAUA Mayflor B.
BAYDO Nyzel Jheune Z.

DE
Alcyren Joy D.
GUZMAN

DE
Giselle Jane C.
GUZMAN
ENCISO Slinnie C.
PROPONENTS
LANDRITO Sofia Lubelle P.
LORIO Alliah B.
NITUDA Jasmine Claire M.
SANTOS Mernelle Mae O.

Last name First Name Middle Initial

Grade Level and Section: 12-STEM 2 Group Number: 1

The Antiseptic Properties of Tinospora cordifolia

Research Title:
(Makabuhay Plant) for Cleansing Hands

N
BIBLIOGRAPHY S OURCE SUMMARY OF CONCEPTS
o.

Mishra,A., Kumar,S. and Research

1 Pandey,A. (2013). Journal Main objectives/background
Scientific Validation of the

48
Medicinal Efficacy
of Tinospora cordifolia.The
Scientific World Journal
Volume 2013, Article
ID 292934
This study aims to identify the scientific evaluation
of Tinospora cordifolia for its medicinal efficacy
which includes phytochemical screening,
antimicrobial, antioxidant, and anticancer activities
of the plant. Secondary metabolites including
anthraquinones, terpenoids, and saponins were
present in many extracts in addition to phenolic.
Research sample/materials used
The Tinospora cordifolia stem was shade-dried,
crushed, and ground into fine powder with mortar
and pestle. Powdered material was sequentially
extracted with petroleum ether (PE), benzene (BZ),
chloroform (CH), ethyl acetate (EA), acetone (AC),
ethyl alcohol (ET), and water (AQ) in Soxhlet
apparatus. The respective extract fractions were
centrifuged, filtered, and lyophilized. The dried
residues were dissolved in DMSO for determination
of antibacterial, antioxidant, and anticancer
activities.
Research method/process
The researchers used the Kirby-Bauer disc diffusion
method which uses inoculum suspension of bacterial
strains that was swabbed on the entire surface of
Mueller-Hinton agar (MHA). Sterile 6 mm diameter
paper discs (Himedia) saturated with 20 L of extracts
prepared in DMSO (containing 3.33 to 10 mg
extract/disc) were aseptically placed on the upper
layer of the inoculated MHA surfaces and plates
were incubated at 37°C for 24 hours. Antibacterial
activity was determined by measuring diameter of
the zone of inhibition (ZOI) surrounding discs.
Main findings/conclusions/recommendations
The study demonstrated the presence of various
groups of phytochemicals in Tinospora
cordifolia extracts which are responsible for showing
considerable antibacterial, antioxidant, and
anticancer activities.
49

Nagaprashanthi,C., Rafi
khan.P, Gopi chand.K,
Aleemuddin MA,and Rajiya
Begum.G. (2012).
In vitro Antimicrobial
Activity of Tinospora
cordifolia and its Research
2 Journal
Phytochemical screening.
International Journal of
PharmTech Research Vol.4,
No.3, pp 1004-1008
Main objectives/background
This study was carried out to evaluate the in vitro
antifungal and antibacterial activity of hydro
alcoholic extract of Tinospora cordifolia creeped on
Azadirachta indica Tree (TC1) in comparison with
that of Tinospora cordifolia (TC2) creeped on
fencing.
Research sample/materials used
The plant material (matured stems) was collected and
shade dried. These dried stems then crushed and
powdered by using electrical grinder. The powdered
material was extracted by hydro alcoholic using
maceration method. The coarsely powdered plant
material was placed in a stoppered container with the
solvent and was allowed to stand at room
temperature for 14 days with frequent agitation until
the soluble matter was dissolved. The mixture was
strained, the marc (the damp solid material) is
pressed, and the extract was concentrated using
distilled evaporator.
Research method/process
In this study the test microorganisms used (bacteria:
Staphylococcus aureus, Pseudomonas aeruginosa,
Escherichia coli and Klebsiella pneumonia; fungi:
Aspergillus niger, Aspergillus fumigates, mucor sp
and Penicillum sp) were obtained from the culture
collections of Department of Microbiology, PRIST
University, Thanjavur, Tamil Nadu, India. The
bacterial were first isolated and subcultured in a
nutrient broth (Oxoid) and incubated at 37°C for 24
h while the fungal were isolated and subcultured on
a Rose bengal agar (RBA) for 72 h at 25°C.
Main findings/conclusions/recommendations
In conclusion from this study, Tinospora cordifolia
stem extract creping on Azadirachta indica has
potential antimicrobial activity similar to that of
neem tree when compared to Tinospora cordifolia
creping on fencing. This can explain that the host
50

Patil,P., Kulkarni,C. and
Pandey,A. (2017).
Antibacterial and
phytochemical analysis of
Tinospora cordifolia,
Azarchita indica and
Ocimum santum leaves
3 extract against
common human pathogens:
An in vitro study Journal of
Pharmacognosy and
Phytochemmistry Volume
6(5): pp 702-706
plant will gain some of the activities when they
survive on medicinal plants.
Main objectives/background
This study aims to identify antibacterial activities of
the leaves of Tinospora cordifolia, Azarchita indica
and Ocimum santum. To prove the validity of
traditional medicine the present work has been
undertaken to evaluate the antimicrobial screening of
leaves of Tinospora cordifolia, Azarchita indica and
Ocimum santum against the human bacterial
pathogens.
Research sample/materials used
Solvent extraction method Thirty grams of dried
powder of Tinospora cordifolia, Azarchita indica
and Ocimum santum leaves were extracted with
aqueous, ethanol, methanol and acetone using
soxhlet apparatus for 48 hrs. The collected extracts
were filtered with Whatman No.1 filter paper and
used for estimation of phytochemicals and
antibacterial activity.
Research
Journal
Research method/process
The modified paper disc diffusion method was
employed to determine the antibacterial activity of
aqueous, ethanol, methanol and acetone extracts.
Turbidity of inoculums was matched with McFarland
turbidity standard. Inoculums were spread over the
Nutrient agar plate using a sterile cotton swab in
order to get a uniform microbial growth. Then the
prepared antibacterial disc were placed over the lawn
and pressed slightly along with positive and negative
controls. Ampicillin 10 mcg/disc were used as
positive control while disc soaked in various organic
solvents and dried were placed on lawns as negative
control. The plates were incubated for 18h at 370 C.
Main findings/conclusions/recommendations
The results obtained in this study thus suggests that
the identified phytochemicals may be the bioactive
constituents responsible for the efficacy of leaves
51

Mamta K. & Jakhar K. (2016)
Veterinary Research
4 International
April-June, 2016 | Vol 4 |
Issue 2 | pp 74-77
extract of Tinospora cordifolia, Azarchita indica
and Ocimum santum against fever, syphilitic, ulcer,
inflammatory disease wounds, conjunctivitis etc.
Based on this, it suggested that the traditional
medicinal use of Tinospora cordifolia, Azarchita
indica and Ocimum santum be continued and
scientific evaluation of its active constituents given
serious attention
Main objectives/background
The present study aims to investigate the in vitro
antibacterial activity of Tinospora cordifolia stem
extract against Escherichia coli O78.
Research sample/materials used
Tinospora cordifolia stems were collected from
Medicinal and Aromatic Plants Section, Department
of Genetics and Plant Breeding, College of
Agriculture, CCS HAU, Hisar.
Research method/process
The stem pieces were washed, shade dried and
powdered in a mixer. For preparation of extract, 15 g
Research of powdered material was boiled in 100 ml of water
Journal and filtered through Whatman`s No.1 filter paper.
The extract was used for assessment of in vitro
antibacterial activity of Tinospora cordifolia stem
extract against Escherichia coli O78.
Main findings/conclusions/recommendations
The overall results of in vitro antibacterial activity of
aqueous extracts of Tinospora cordifolia against E.
coli indicated that the extract has potential to be used
commercially for control of E. coli infection. But
further in vivo trials on domestic animals and poultry
infected with E. coli need to be conducted to evaluate
its clinical efficacy before recommending for field
use on large scale. However, it can be concluded that
in the light of increased antibiotic resistance, the
present investigation is of importance for preparing
Tinospora cordifolia based antimicrobials.
52

Stuart Jr.,G.(2018).Tinospora
crispa (L.) Hoook.f. &
5 Online Source
Thomson Heavenly Elixir.
Philippine Medicinal Plants
Mohammed,A.,Manish,G.,&D
inesh(2012).Antimicrobial
Activity of Tinospora crispa
Root Extracts. Research
6 Journal
International Journal of
Research in Ayurveda and
Pharmacy. Vol 3(3)
Main points/concepts
The Tinospora codifolia has wide range of use in
terms of its ability as an alternative medicine in
which the stems and leaves of the plant are the main
parts that are utilized. Its properties are considered
febrifuge, vulnerary, tonic, antimalarial,
parasiticide, and insecticidal. Studies also suggests
Tinospora cordifolia effective as an antioxidant,
antidiarrheal, antimicrobial, anti-diabetic, anti-
proliferative, cardiotonic, hypolipidemic,
antimalarial, antinociceptive properties. Even
though it has many uses, there are cautions in using
the plant especially those who are pregnant or
patients with cardiac disorders.
Main objectives/background
This study aims to determine the in vitro
antimicrobial activity of ethanol, distilled water,
methanol, and chloroform crude extracts of
Tinospora crispa. Antimicrobial activity will be
examined against gram positive bacterial strains.
And more specifically, the maximum and minimum
zone of inhibition against the bacterial strains.
Research sample/materials used
Tinospora crispa roots were collected around the
area in Selangor, Malaysia then grounded for this
study. Also 3 microorganisms were used namely
Stretococcus pneumonia (S.pneumonia), Escherichia
coli (E. colli), and Candida albicans (C.albicans).
The rationale behind choosing these microorganisms
against T.crispa roots is that one of the objectives of
this study was to examine the antimicrobial activity
of the root extract against gram positive bacterial
strains.
Research method/process
The study used the extraction method to get the
ethanol, distilled water, methanol, and chloroform
crude extracts of Tinospora crispa. This uses disc
diffusion method against standard antibiotic discs
53

Aminul,H.,Ashraful,I., and
Mohammad,S. (2011).
Antimicrobial,
Cytotoxicity,and Antioxidant
activity of Tinospora
7 cordifolia. Journal of
Pharmaceutical and
Biomedical Sciences. Vol
13(12)
which were used for comparison then the autoclaving
method was used to sterilize the media in which the
microorganisms were put into. The bacterial culture
was then swabbed onto the medium in petri dishes
and disc impregnated with extracts were placed on
sold agar medium then placed in an incubator.
Main findings/conclusions/recommendations
Initial results or findings showed that even though
the plant’s ethanol and chloroform extracts have
provided the best results as an antimicrobial this
only showed the basic information on the
antimicrobial potential of the plant so further studies
are required to substantiate new biologically potent
active antimicrobial compounds.
Main objectives/background
This study aims to determine the antimicrobial,
cytotoxicity, and antioxidant activity of Tinospora
cordifolia. The antimicrobial, cytotoxicity, and
antioxidant activity was examined through the
extracts of the plant that was subjected to
antimicrobial screening against some gram-positive
and gram-negative organisms.
Research sample/materials used
The plant sample of Tinospora cordifolia was
Research
Journal collected from Meymensing which were sun dried
and grounded into course powder and this will be
then used against some gram-positive and gram-
negative bacteria. The reason of selecting these
materials are to determine the antimicrobial,
cytotoxicity, and antioxidant activity of Tinospora
cordifolia.
Research method/process
The study used the extraction and isolation method
to get the petroleum ether, methanol, chloroform,
carbon tetrachloride, aqueous soluble fractions of
extracts of Tinospora crispa. Antimicrobial
54

Razo et al (2015).
Antibacterial potential of
Makabuhay (Tinospora
8 Journal
cordifolia) Extract
formulated as Facial Cleanser
screening is used specifically the disc diffusion in
measuring the diameter of zone of inhibition. The
brine shrimp lethality bioassay technique was used to
determine the cytotoxicity of the plant extractives.
And the method developed by Brand-Williams et al
was used to determine the antioxidant activity or the
free radical scavenging activity.
Main findings/conclusions/recommendations
Initial results or findings indicated that the extracts
that were collected possess antioxidant, cytotoxic,
and few antimicrobial substances which justify the
traditional uses of this plant in the treatment of
different diseases. Furthermore more future
researches are necessary for elucidating the active
principles.
Main objectives/background
This study aimed to determine the efficacy of
Makabuhay plant extract which was formulated as a
facial cleanser subjected to different concentrations
of the plant extract (50%, 75%, and 100%
concentrations) since Makabuhay (Tinospora
cordifolia) plant has been identified to be having
antibacterial properties, giving rise to the
formulated products such as ointments and soaps
were which are now commercially available.
Research sample/materials used
Research
Tinospora cordifolia extracts were collected and
formulated as a facial cleanser and Staphylococcus
aureus is the specific bacteria which was used since
it is a causative agent of pimple and acne.
Research method/process
Kirby-Bauer Test/ Disk-diffusion method was used
to determine the antibacterial activity of Makabuhay
plant extract after preparing assay plates for five
trials. The test for the efficacy of the formulated
facial cleanser against the control was determined by
susceptibility testing using disk-diffusion method. A
zone of inhibition (mm) was measured for every trial
55

Archa Vermani, Navneet and
Shiv Shanker Gautam, (2013).
Screening of Antibacterial
Activity of Tinospora
cordifolia Miers. Extracts
9 Against Dental Pathogens.
Journal of Pharmacology and
Toxicology, 8: 28-34. DOI
10.3923/jpt.2013.28.3410.392
3/jpt.2013.28.34
at different concentrations and was compared with
the control facial cleanser.
Main findings/conclusions/recommendations
Initial results or findings indicated that the
formulated Makabuhay facial cleanser showed
significant antibacterial activity against
Staphylococcus aureus at 50% concentration, 75%
concentration, and 100% concentration as the most
effective. The control particularly showed
insignificant activity against Staphylococcus
aureus. The experiment confirms the antibacterial
property of Makabuhay (Tinospora cordifolia) plant
extract as a formulated facial cleanser against
Staphylococcus aureus at different concentrations.
Main objectives/background
This study aims to investigate the crude extracts of
Tinospora cordifolia’s ability to inhibit the growth of
dental pathogens such as Staphylococcus aureus,
Streptococcus mutans, Streptococcus salivarius,
Lactobacillus acidophilus, Streptococcus sanguinis.
Other than that, it also aims to search an alternative
way for managing such infectious diseases with
efficient cure and very low or no side effects.
Research sample/materials used
Research The researchers used the stems of Tinospora
Journal
cordifolia local market, Haridwar and authenticated
at Botanical Survey of India, Northern Regional
Center, Dehradun to test its antibacterial
performance to dental pathogens.
Research method/process
The methods used are as follows:
(1) Preparation of the extract by immersing 200 gm
of dried powdered material in 600 mL of solvents.
(2) Test microorganisms where bacterial strains were
grown and maintained on nutrient agar slants at 4°C.
56

Sankhala, L.N., Saini,R.K.
and Saini, B.S. (2010) A
Review on Chemical and
Biological
10 Properties of Tinospora
cordifolia Vol. 2, No. 2, pp.
340-344., Int. J. Med.
Arom. Plants, ISSN 2249 –
4340
11 Patil,P., Kulkarni,C. &
Pandey,A. (2017)
(3) Antibacterial testing, where disk diffusion
method was used and that to test the extent of the
growth of added microorganism in a zone around the
hole containing a solution of tested material.
(4) Test for alkaloids, flavonoids, glycosides,
steroids,saponins, tannins
Main findings/conclusions/recommendations
It was found that the methanol property of the extract
was the most effective against all tested bacterial
pathogens. Therefore it is concluded that the stems of
T. cordifolia has a good antibacterial activity and its
properties help in inhibiting the growth of
microorganisms.
Main points/concepts
This article aims to determine and assess the
chemical and biological properties of Tinospora
cordifolia.
Tinospora cordifolia’s chemical constituents were
alkaloids, glycosides, diterpenoid, lactones,
sesquiterpenoids and steroids. Along with that, the
plant also contains a lot of biological properties such
as that it has been reported that the extract of T.
cordifolia has free radical scavenging and
antioxidant effect. It has seen effective in alloxan
induced diabetic rats which shows an increase in the
Article concentration of TBARS in kidney and liver of
diabetic rats. It also shows hypoglycemic effect that
the oral administration of the aqueous extract of the
plant caused to reduce the blood glucose and brain
lipids of the rats. The aqueous and alcoholic property
of the plant also help improve the immuno-
modulatory activity of an organism and the
modulation of hepatic functions. This is to conclude
that various type of studies regarding T. cordifolia
shows its effectiveness as a remedy for various
ailments.
Research Main objectives/background
Journal
57
 Antifungal and
phytochemical properties of
Tinospora cordifolia,
Azadirachta indica and
Ocimum sanctum leaves
extract. Journal of
Medicinal Plants Studies
2017 5(5): 23-26
Duraipandiyan,V.,
12 Ignacimuthu,S., and
Balakrishna,K. (2012)
Antimicrobial Activity of
Tinospora cordifolia: An
Ethnomedicinal Plant.
This study aims to determine the antifungal activity
of the plants T. cordifolia, A. indica and O. sanctum
that would later then lead to the establishment of
compounds that could be used to formulate new
antimicrobial drugs of natural origin.
Research sample/materials used
The presence of the aqueous, methanol and acetone
of T. cordifolia, A. indica and O. sanctum in the
extract of the leaves were tested for the presence of
phytochemical components against Candida
albicans, Aspergillus niger, Aspergillus fumigatus,
Microsporum gypseum and Trichophyton rubrum.
Research method/process
The process includes the preparation of the plant
material by drying and powdering and the
preparation of the extracts using the filter paper.
Then the phytochemical screening which includes
the test for tannins, saponins, steroids, glycosides
and anthraquinones. The method of disk diffusion
was also used to determine the antifungal activity of
solvent extract of leaves.
Main findings/conclusions/recommendations
T. Cordifolia shows to have more potential against
microbial infections. It contains more of the active
compounds and a high potency where it showed
antifungal activity among the entire fungal
organism.
Main objectives/background
This study aims and searches for natural products
that has an effect in the antimicrobial activity, one of
them is the known medicinal herb in India, Tinospora
Research
Journal cordifolia.
Research sample/materials used
58
 Retrieved from:
https://www.yumpu.com/en
/document/view/26663609/
antimicrobial-activity-of-
tinospora-cordifolia-an-
ethnomedicinal-plant
Singh, K., Panghal, M.,
Kadyan, S., Chaudhary, U., &
Yadav, J. (2014). Antibacterial
Activity of Synthesized Silver
Nanoparticles from Tinospora
cordifolia against Multi Drug
Resistant Strains of
Pseudomonas aeruginosa
Isolated from Burn Patients.
13 Retrieved from
https://www.omicsonline.org/o
pen-access/antibacterial-
activity-synthesized-silver-
nanoparticles-tinospora-
cordifolia-multi-drug-
resistant-strains-pseudomonas-
aeruginosa-isolated-burn-
patients-2157-7439-5-
192.php?aid=25859
The materials used were the extracts of Tinospora
cordifolia and the bacteria Bacillus subtilis,
Enterococcus faecalis, Staphylococcus aureus,
Escherichia coli, Klebsiella pneumoniae, Proteus
vulgaris. Fungi such as T. rubrum, T.
Mentagrophytes, T. simii, Epidermophyton
floccosum.
Research method/process
The research method includes the collection of the
plant material along with the preparation of crude
extracts. They also isolated and identified the active
compound and test organism such as Bacillus subtilis
and Enteroccocus faecalis. Then the disc diffusion
method and lastly, the antifungal activity and
minimum inhibitory concentration were performed
accordingly.
Main findings/conclusions/recommendations
The antimicrobial screening results indicated that
both antibacterial and antifungal activities were
observed in the ethanol extract of the plant.
Main objectives/background
This study is based on the synthesis of silver
nanoparticles from Tinospora cordifolia and checked
its antibacterial activity against MDR strains of P.
aeruginosa.
Research sample/materials used
Research
Journal The stem of Tinospora cordifolia was collected locally
from Botanical Garden, M.D. University, Rohtak,
Haryana, India. It was thoroughly washed in distilled
water, cut into fine pieces. 15 g of fresh plant material
was boiled into 100 ml sterile distilled water and
filtered through Whatman`s No.1 filter paper. The
extract was stored at 4°C for further experiments. The
aqueous solution of 1 mM silver nitrate (AgNO 3 ) was
prepared and used for the synthesis of silver
nanoparticles. 15 ml of plant extract was added into 200
ml of aqueous solution of 1 mM silver nitrate for
59

Patel, M., & Ishnava, K.
(2014). In vitro Study of
14 Tinospora Cordifolia (Wild.)
Miers (Menispermacceae)- a
Multipurpose Plant, by Using
Different Plant Bark Extracts
reduction into Ag+ ions and kept for 15-20 minutes at
70-75°C. This aqueous extract acts as reducing and
stabilizing agent for 1mM of AgNO3 . The prepared
AgNPs were further characterized
Research method/process
The techniques used, first is UV-VIS spectroscopy
wherein the Ag nanoparticles were characterized in a
Shimadzu UV-VIS Spectrophotometer. The Fourier
Transform Infra-red Spectroscopy (FTIR), To
remove any free biomass residue or compound that is
not the capping ligand of the nanoparticles, after
complete reduction, silver nanoparticles were
concentrated by repeated centrifugation (3 times) of
the reaction mixture at 15,000 rpm for 20 min. Third,
Transmission Electron Microscopy (TEM) and
Energy Dispersive X-Ray Spectroscopy (EDX), the
shape and size of AgNPs was determined by
transmission electron microscopy. The X- Ray
Diffraction (XRD), The metal nanoparticles were
coated on to the glass substrate and after drying the
sample was analysed by X-ray diffractometer. And
lastly, Multi drug resistant P. aeruginosa from
clinical isolates, Twenty P. aeruginosa isolates were
obtained from the various samples of burn patients
receiving in Microbiology Department of Post
Graduate Institute of Medical Sciences, Rohtak.
Main findings/conclusions/recommendations
From the study they conclude that even at very small
concentration (in μg/ml) AgNPs from stem of T.
cordifolia possess very good antibacterial activity
which makes them a potent source of antibacterial
agent against MDR strains of P. aeruginosa. Also,
green synthesis of AgNPs can potentially eliminate the
problem of chemical agents that may have adverse
effects, thus making nanoparticles more compatible
with the eco-friendly approach. Moreover the
synthesized AgNPs enhance the therapeutic efficacy
and strengthen the medicinal values of T. cordifolia.
Main objectives/background
The study was undertaken to increase secondary
Research
Journal
metabolite production by adding plants extracts in
plant tissue culture medium.
60
for Secondary Metabolite Research sample/materials used
Production. Journal of Herbs,
Mature plants of T. cordifolia were collected and
Spices and Medicinal Plants,
bark of Azadirachta indica (Neem) and Acacia
341-349.
nilotica (Baval) trees from the campus of New
doi:10.1080/10496475.2013.8
Vallabh Vidyanagar. The plants were identified at
76482
the Ashok and Rita Patel Institute of Integrated
Study and Research in Biotechnology and Allied
Sciences, New Vallabh Vidyanagar, Gujarat, India.
Murashige and Skoog medium with sucrose 3%,
0.8% agar, and growth hormones (2, 4-D) were
used. The medium was prepared with combination
(1.0 mg.L−1 , 1.5 mg.L−1 , and 2.0 mg.L−1 of Neem
and Baval bark extracts. The concentrated stock
solutions of ingredients were prepared and
refrigerated.

Research method/process
The surface sterilization of explants (leaves and
nodal segments) was
carried out as per as stranded protocol with slight
modifications to
include immersion of explants in 0.1% mercuric
chloride (HgCl2 ) solution for 4 min. Explants were
inoculated and experimental manipulations carried
out under strictly aseptic conditions in a laminar air
flow bench.
The tubes and bottles were shifted to a culture room
with controlled facility of diffused light (2,000 lux)
for 10 h daily at 28 •} 2◦ C and 50 to 60% relative
humidity. The calluses were collected after 30 to 40
days, washed with distilled water to remove all
adhering particles, and dried at room temperature.
Callus extracts were prepared as described earlier,
evaporated, and dissolved in methanol for further
analyses.

Main findings/conclusions/recommendations
Leaves of T. cordifolia are rich in protein (11.2%)
and are fairly rich in calcium and phosphorus and
containing anti-oxidant activity in vitro models. The
use of T. cordifolia for the treatment of general
weakness and fever, the immunomodulatory

61

Saha S. & Ghosh S., (2012).
Tinospora cordifolia: One
plant, many roles. Retrieved
from:
15 https://www.ncbi.nlm.nih.g
ov/pubmed/23661861
16 Sharma,V. & Pandey,V .(
2010). Beneficial Effects
properties, hypoglycemic activities have been
reported. Three major groups of compounds—
protoberberine alkaloids, terpenoids, and
polysaccharides—are considered as putative active
constituents of T. cordifolia. Protoberberine
alkaloids such as berberine and palmatine are
reported to have anti-cancer, anti-infective, anti-
diabetic and immunomodulatory activities.
Main objectives/background
This study aims to show the importance of the
effectiveness of plant genetic resources that can help
to prevent or to lessen the bacteria that can be the
cause of the illness in our body.
Research sample/materials used
The researchers used information extracted from a
total of 175 published article of which five review
articles and cross references thereof were collected.
The search criteria were restricted to the roles of the
plant in the field of medical advancements and the
effects that has been observed with different
experiments.
Research
Journal
Research method/process
The study was conducted through vivo analysis. The
reports of morphological and physiological
characters of the plant in which it is on the recent
development published literature including original
articles and papers Pubmed and Pubmed Central
Databases were taken into study for the report.
Main findings/conclusions/recommendations
The study shows that the plant has the versatile
resource for all forms of life. All the active
compounds that are conducted have roles of different
types. And the Tinospora cordifolia is enabling
effectiveness disease targeting. The plant is truly
helpful and an incredible source for world medicine.
Research Main objectives/background
Journal
62
 of Tinospora cordifolia on This study aims to prove that the stem and the leaves
Blood Profiles in Male do not just apply or treats for humans but also for
Mice Exposed to Lead. animals. But it is widely used for humans because of
Retrived from the bacteria and the diseases that most people
encounter.
https://www.ncbi.nlm.nih.g
ov/pmc
/articles/PMC2964740/ Research sample/materials used
The researchers used chemicals and Tinospora
cordifolia was collected from the medical garden of
Banasthali University, Rajasthan, India, and
identified by a plant taxonomist of the institute. The
plant materials (stem and leaves) were thoroughly
washed with distilled water and by using the
laboratory homogenizer. The shade dried and cut into
small pieces, and powdered separately. And also the
mice that were used was weighing approximately 15-
30g. Known quantities of the powdered materials
were extracted separately using distilled water as a
solvent. The extracts were then filtered through filter
paper and concentrated on water bath. And after
complete evaporation of the solvent, the residue were
weighed and stored at 4°C and used to treat the
animals as needed.

Research method/process
The study was conducted through ANOVA which is
a statistical technique that assesses potential
differences in a scale-level dependent variable by
nominal-level variable having 2 or more categories.

Main findings/conclusions/recommendations
The study shows that the stem and leaves crude
extracts are capable of scavenging lead-induced
hematological alternations to some extent and stem
and leaf extract was observed leaf extract were also
observed as compared to normal animals. Thus,
proves that the plant can be both used in humans
and animals.

Kavya B, (2015). Tinospora Main points/concepts

17 Cordifolia (WILLD.) MIERS.: Online Source
This study aims to show the various benefits
Nutritional,Ethnomedical and acquired by the use of Tinospora cordifolia as

63
 Therapeutic Utility. Retrieved
from
http://www.ijrap.net/admin/ph
p/uploads/1326_pdf.pdf
Singh, P., Katiyar, D., Singh,
B. & Srivastava, A. (2015).
Antimicrobial activity of
Tinospora cordifolia extracts
18
against urinary tract infections
causing bacteria. International
Journal of Pharma and Bio
Sciences. 6. B571-B577.
Rekha, Tamilarasu S., & Viji
19 (2017) Tinospora cordifolia.
Retrieved from
treating various ailments. The researchers used the
whole plant, which is the powdered root, stem bark
and the decoction of root and stem. This study
shows that even though there are many herbal plant
medicines in the world, the Tinopora cordifolia has
a higher medical value. Because of the stem and
leaves, it is used for treating various ailments.
Main objectives/background
This study aims to test the antimicrobial property of
Tinospora cordifolia extract against urinary tract
infections causing bacteria. Antimicrobial activity
will be examined against gram positive and gram
negative strains.
Research sample/materials used
The researchers mainly used a sample of Tinospora
cordifolia. The plant sample was thoroughly washed
with water then air-dried sample was subjected to
drying at an oven with a temperature of 37 degrees
celcius. Dried stem was crushed by hand then was
crushed in a mixer grinder to coarse powder.
Research
Journal
Research method/process
The study used the solvent extraction method to get
the supernatant and increases the volume of the
solvent one fourth than the original one. They used
the disk diffusion method in measuring the diameter
of the inhibition zones.
Main findings/conclusions/recommendations
Initial results or findings showed that Tinospora
cordifolia can be a good and potential source of
herbal drug preparation for treating UTI’s. The active
principles were more soluble in ethanol than
methanol. The researchers recommended that by
increasing the concentration of extracts its activity
can also increase further.
Main points/concepts
Online Source
Tinospora cordifolia can be seen on subtropical and
tropical countries. Tinospora cordifolia has a lot of
64
 http://vikaspedia.in/agriculture
/crop-production/package-of-
practices/medicinal-and-
aromatic-plants/tinospora-
cordifolia
Choudhary, V. (2018)
Description and medicinal
uses of Guduchi – Tinospora
cordifolia.
20 Retrieved from
http://natureconservation.in/de
scription-and-medicinal-uses-
of-guduchi tinospora-
cordifolia
Hook, F. & Thomas (2018)
Uses of Tinospora cordifolia
21 (Willd.) Retrieved from
http://nif.org.in/TINOSPORA-
CORDIFOLIA
Joshi,G. and Kaur,R.
(2017). Tinospora
cordifolia : A
Phytopharmacological
Review. International
Journal of Pharmaceutical
22
Sciences and Research
http://ijpsr.com/bft-
article/tinospora-cordifolia-
a-
phytopharmacologicalrevie
w/?view=fulltext
theraputic uses. Its stem contains bitter substances. It
has other compounds such as gilonin, gilosterol,
gilenin, etc. The plant according to them is more
effective than a dried one. It has an alterative, diuretic
and aphrodisiac properties. It can be used in febrifuge
used in chronic fever. The plant Tinospora cordifolia
can cure fever, urinary disorders rheumatism and
dyspepsia.
Main points/concepts
Tinospora cordifolia grows in deciduous and dry
forests. It has many medicinal properties like
antibiotic, immunosuppressant, anti-cancer, anti-
spasmodic, anti-microbial, anti-osteoporotic, anti-
inflammatory, anti-arthritic, anti-allergic, anti-
Online Source diabetic, anti-oxidant, anti-stress etc. A variety of
active components like alkaloids, steroids,
diterpenoid lactones, aliphatics, and glycosides,
have been isolated from the different parts of the
plant body, including the root, stem, and leaves of
the plant. Guduchi is a traditional Ayurvedic
medicine, and used in the treatment of fever,
diabetes, diarrhea, cancer, pain, skin disease etc.
Main points/concepts
Tinospora cordifolia is a herbal medicine that has a
lot of use. It is a well known medicinal plant and
Online Source used to cure a number of disease. It is also called as
magical herb due to its property of curing a lot of
diseases. It is commonly used to cure migraine,
asthma, fever, jaundice, diabetes, intestinal worms
and piles.
Main objectives/background
This study aims to prove that the Tinospora
cordifolia has been one of the important sources of
medicines since beginning of human civilization and
to present the various parts and their actions.
Research
Although the review articles on this plant are already
Journal published, this review article is presented to comply
all the updated information on its phytochemical and
pharmacological activities, which were performed by
widely different methods.
Research sample/materials used
65
 The researchers used the traditional form of a plant
which is the Tinospora cordifolia that is native to
India and widely used from ancient medical history.

Research method/process
The study was conducted by investigating and a year
of experimenting.

Main findings/conclusions/recommendations
This study shows that even though there are many
herbal plant medicines in the world, the Tinospora
cordifolia has a higher medical value. Because of the
stem and leaves, it is used for treating various
ailments.

Main points/concept
The objective of this review is to extend the current
knowledge, importance and beneficial
pharmacological applications of T. cordifolia in
Dadar, M., et al
humans for safeguarding various health issues. The
(2017).Medicinal and
present review describes medicinal applications of
Beneficial Health T. cordifolia in countering various disorders and
Applications of Tinospora usages as anti-oxidant, anti-hyperglycemic,
cordifolia (Guduchi): A antihyperlipidemic, hepatoprotective,
Miraculous Herb cardiovascular protective, neuroprotective,
Countering Various osteoprotective, radioprotective, anti-anxiety,
Diseases/Disorders and its adaptogenic agent, analgesic, anti-inflammatory,
antipyretic, a thrombolytic agent, anti-diarrheal,
23 Immunomodulatory Effects. Online Source
Recent Pat Endocrine, anti-ulcer, anti-microbial and anti-cancer agent. The
Metabolic and Immune plant is also a source of micronutrients viz. copper,
Drug Discovery, 92- calcium, phosphorus, iron, zinc and manganese. A
special focus has been made on its health benefits in
111.doi:10.2174/187221481 treating endocrine and metabolic disorders and its
1666170301105101 potential as an immune booster. Several patents
have been filed and granted to inventions
encompassing T. cordifolia as a major component
of therapeutics for ameliorating metabolic,
endocrinal and several other ailments, aiding in the
betterment of human life expectancy. The
information presented would be beneficial for
researchers, medical professionals and
pharmaceutical companies to design and develop

66

Hipol R. L., Cariaga M. F. N.
M., Hipol R. M., (2012) .
Anti-Inflammatory Activities
of the aqueous extract of the
stem of Tinospora crispa
24 (family menispermaceae).
Retrieved from:
http://www.journalofnaturestu
dies.org/files/88-95-
Tinospora-full.pdf
effective medicines, drugs and healthical products
exploiting the multiple as well as specific modes of
actions of T. cordifolia, and also help in promoting
and popularizing this rich herb having promising
potentials to prevent and treat various ailments.
Main objectives/background
The study aims to evaluate the anti-inflammatory
properties of the aqueous extract of the dried stem of
the plant in order to provide scientific grounding to
its use.
Research sample/materials used
The researchers used the stems of Tinospora crispa
that was collected from Cabalayangan, Bauang, La
Union. Plant extraction from the dried stem powder
to fill the thimble for Soxhlet extraction was weighed
in a digital balance. Using distilled water as solvent,
the stems were subjected to Soxhlet extraction.
About Forty five albino rats of either sex weighing
180-200g were used for the Carageenan-induced paw
edema assay. For the Membrane Stabilization test,
they took a healthy blood samples from the Clinical
Research Laboratory of the College of Natural Sciences, Saint
Journal Louis University.
Research method/process
The study used experimental method wherein they
observe and test the albino rats induced with acute
inflammatory using the methods by Lawal et al. The
rats were treated with different concentrations of
Tinospora crispa extracts with different
concentrations and observed in succeeding hours
computing it by the said formula. The statistical
Analysis of the data were subjected to a One-Way
ANOVA at p< 0.05, using SPSS program.
Main findings/conclusions/recommendations
The study shows that the anti-inflammatory
properties of the aqueous extract were induced to
albino rats on different doses (50 mg/kg, 100 mg/kg
and 150 mg/kg) it revealed that it reduced the paw
67

Bala M, Pratap K, Verma PK,
Singh B, Padwad Y.
Validation of ethnomedicinal
potential of Tinospora
cordifolia for anticancer and
immunomodulatory activities
and quantification of bioactive
25
molecules by HPTLC. J
Ethnopharmacol. 2015 Dec
4;175:131-7. doi:
10.1016/j.jep.2015.08.001.
Epub 2015 Aug 5. PubMed
PMID:
26253577.
edema in rats and the protein stabilization test had the
same effect as ibuprofen in preventing protein
denaturation. Thus, proves that the plant may used to
treat inflammation disorders.
Main objectives/background
The study aims isolation, structure elucidation,
quantification and pharmacological evaluation of
secondary metabolites from Tinospora cordifolia for
anticancer and immunomodulatory activities.
Research sample/materials used
A fraction of different extracts of the stem of
Tinospora cordifolia were used.
Using it against to four different human cancer cell
lines such as KB (human oral squamous carcinoma),
CHOK-1 (hamster ovary), HT-29 (human colon
cancer) and SiHa (human cervical cancer).
Research method/process
Research The study used experimentation method, whereas
Journal
pure molecules were isolated using normal phase
chromatography and characterized based of NMR
and mass spectroscopic techniques. The anti-cancer
and immunomodulatory activities of different
extracts, fractions and isolated compounds were
evaluated against four different human cancer cell.
Using HPTLC method was also developed for the
quantification of three bioactive compounds i.e N-
formylannonain (1), 11-hydroxymustakone (5) and
yangambin (8) in the stem of T. cordifolia hosted on
fifteen different plants.
Main findings/conclusions/recommendations
There are a total of eight compounds that was
isolated and characterized to different classes. The
pharmacological evaluation of extract, fractions and
pure molecules revealed the ethnomedicinal value of
T. cordifolia for anticancer and immunomodulatory
activities.
68

Kumar DV, Geethanjali B,
Avinash KO, Kumar JR,
Chandrashekrappa GK et al.
(2017) Tinospora cordifolia:
The Antimicrobial Property of
26
the Leaves of Amruthaballi. J
Bacteriol Mycol Open Access
5(5): 00147. DOI:
10.15406/jbmoa.2017.05.0014
7 extract the leaves by washing the climbers leave and
Main objectives/background
The study aims to know The antimicrobial property
of the leaves of Tinospora cordifolia. They used
the said plant interest the researcher since the
medicinal properties are widely popular in the
Ayurveda medicine. The leaves were tested and
observed on Escherichia coli on three different
solvent. Ethanolic, methanolic and aqueous
extracts of the leaf were tested using slip disc
method to amount the anti-microbial againts E.
coli.
Research sample/materials used
The research used Tinospora cordifolia leaves were
collected from Dixit clinical from Vijaynagar in
Mysore, Karnataka and the pathogen, Escherichia
coli (E. coli) a Gram negative rod aerophilic
bacterium was collected from JSS Medical College,
Mysore, India (Anil Kumar Sir). The plates obtained
were cultured on Nutrient Agar Medium.
Research
Journal
Research method/process
They use experimentation method wherein they
shade dry them then the dried leaves were powdered
using mortar. The powdered T. cordifolia leaves
were divided into 3 groups each 20g leaf powder.
The groups then subjected to polar and aqueous
extract, the leaf powder was soaked in 50 ml of the
solvents ethanol, methanol and aqueous extract. The
soaked plant extracts then incubated at room
temperature for two days. They then obtain and used
the crude nature of the extract to further test the anti-
microbial nature of the leaf extract against the test
microorganisms.
Main findings/conclusions/recommendations
As the result, the medicinal plant proves that it can
be an alternative form of medicine than the
consumers used with its fewer side effects and easy
availability. The old tradition with the touch of
69

Upadhyay A. K., Kumar K.,
Kumar A., Mishra H.S.,
(2010). Tinospora
cordifolia (Willd.) Hook. f.
27 and Thoms. (Guduchi) –
Research
Journal
validation of the Ayurvedic
pharmacology through
experimental and clinical
studies. Retrived from:
https://www.ncbi.nlm.nih.gov/
pmc/articles/PMC2924974/
modern technology is said to have new hope of
medicine where the base is founded the
phytocompounds from the plants and the alterations
to the mode of delivery are formed by the advanced
technology. The understanding and knowledge of
medicinal plants and the age old practice is the
primary and most important step in achieving the
final result.
Main objectives/background
Tinospora cordifolia (Guduchi) is a widely used plant
in folk and Ayurvedic systems of medicine. Reports
of the chemical constituents of the said plant belong
to different classes, such as alkaloids, diterpenoid
lactones, glycosides, steroids, sesquiterpenoid,
phenolics, aliphatic compounds and polysaccharides.
In ancient texts of Ayurveda, various properties of T.
cordifolia acquires scientific validity through
adopting “reverse pharmacological” approach.
The review will draw together various properties and
medicinal uses of T. cordifolia along with
phytochemical and pharmacological reports.
Research method/process
The research method is systematic literature review
wherein the review will validate the ancient texts of
Ayurveda, like Rasayana, Sangrahi, Balya,
Agnideepana, Tridoshshamaka, Dahnashaka,
Mehnashaka, Kasa-swasahara, Pandunashaka,
Kamla-Kushta-Vataraktanashaka, Jwarhara,
Krimihara, Prameha, Arshnashaka, Kricch-
Hridroganashak.
The review also gives out detailed tables such as
Ayurvedic properties (dravya-guna) of T.
cordifolia (Guduchi), Karma (action -
pharmacodynamics) and prayoga (uses) of T.
cordifolia, Uses of T. cordifolia (Guduchi) in folk
and tribal medicine, and Chemical constituents of T.
cordifolia (Guduchi). Indian contributions to the
therapeutic revolution through reverse
pharmacology will have to integrate throughput
screening, combinatorial chemistry and effects of
the old or novel compounds/ plants on human gene
expression and proteomics.
70
 The study also collects the medicinal properties
reports of the said plant, which includes anti-
diabetic, antipyretic, antispasmodic, anti-
inflammatory, anti-arthritic, antioxidant, anti-
allergic, anti-stress, anti-leprotic, antimalarial,
hepato-protective, immuno-modulatory and anti-
neoplastic activities.

Main findings/conclusions/recommendations
In conclusion, the medicine has great potential in
modern pharmacotherapeutics. A remarkable body
of modern evidences validated the pharmacological
actions attributed to Tinospora cordifolia in
Ayurvedic texts.

Main objectives/background

The study aims to show the links between what is

believed and what is known regarding the use of
Tinospora cordifolia.

Mutalik,M.,& Mutalik N. Research sample/materials used

(2011) Tinospora cordifolia Evidences from pharmacological studies were used
and its Varied Activities: to know the possible mechanisms of its claimed
What is believed benefits.
And What is known?.

International Journal of Research Main findings/conclusions/recommendations

28 Journal
Current Research and
The researchers suggest that Tinospora cordifolia
Review Vol. 03
has antioxidant, radio protective,
Retrieved from antihyperglycemic, anti-inflammatory, anti-ulcer
http://ijcrr.com/uploads/20 and antispasmodic properties, and capacity to
dissolve urinary calculi.
25_pdf.pdf
It is also a potential antidepressant and enhances
cognition and memory. Immunemodulation,
preventing oxidative damage, and relieving
inflammation are likely explanations or benefits in
infections, joint inflammation, and allergies. The
constellation of various
activities plays a role in its protective effects against
antitubercular and cytotoxic drugs, and toxins.

71

Castillo et al. (2013).
Efficacy and safety
of Tinospora cordifolia
lotion in Sarcoptes scabiei
var hominis- infected
pediatric
29 Patients: A single blind,
randomized controlled trial.
Retrieve from
https://www.ncbi.nlm.nih.g
ov/pmc/articles/PMC36433
41/
Arcueno, R., Retumban,J.,
Echao,J.,and
Guerrero,J.(2015).Wound
healing potential of
Tinospora crispa (Wild.)
Miers [Menispermaceae]
stem on diabetic mice.
30
Journal of Medicinal Plants
Studies 2015; 3(2):106-109.
Retrieved from
http://www.plantsjournal.co
m/archives/2015/vol3issue6
/PartB/3-6-8.pdf
Main objectives/background
The researchers evaluated the clinical efficacy and
safety of Tinospora cordifolia lotion including its
cure rate and clearance time compared with
permethrin lotion.
Research method/process
A clinical study was performed in three government
institutions to investigate clinical efficacy
of Tinospora cordifolia lotion in sixty-six scabies-
infected patients.
Main findings/conclusions/recommendations
T. cordifolia lotion and permethrin significantly
reduced the mean global evaluation score after four
weeks of treatment. The two lotions showed
comparable effects as anti-scabies agent. Moreover,
the clearance time (days) and cure rate using the two
lotions did not differ. Clinical improvement, mean
clearance time and cure rate of T. cordifolia lotion
are comparable with permethrin.
Main objectives/background
The researchers aimed to compare the effect of
Tinospora crispa on healing of diabetic wound
among albino mice when given and administered
along with an ointment topically applied to the
wound.
Research Research sample/materials used
Journal
Preparation of Tinospora crispa ointment was done
using a common household procedure in the
Philippines. Fresh stem weighing 100g were cut
into small pieces and slowly fried until crispy in 200
ml of vegetable oil. Stems were removed while the
oil was mixed with two wax candles sliced into
small cubes. All debris were strained. The wax was
thoroughly mixed and allowed to melt and settle.
The researcher also used Albino mice 8-10 weeks
old and about 25-30 grams. The animals were
72
acclimatized for seven days before proceeding with
the experiment.

Research method/process
Numerical data were analyzed using Analysis of
Variance (ANOVA) at 5% level of significance and
Duncan Multiple Range Test (DMRT) as post-hoc
test. Statistical analysis was carried out using SPSS
version 22.

Main findings/conclusions/recommendations
From the results of the study, Tinospora crispa stem
extracts showed potentials to be used as a remedy
for diabetic wounds, primarily in reducing blood
glucose level and second, in possibly preventing
microbial infection in affected area.

73
 Appendix C
DOCUMENTATION

Soaking of powdered stems in ethanol Filtration of the ethanolic extract

Rotary evaporation of ethanolic extract Cooking of the agar

Making of the different concentrations Soaking of disks into diff. concentrations

74
Pouring of agar into petri dishes Swabbing of bacteria (S.aureus) into the agar

Putting of disks into the agar Wrapping and labeling of petri dishes

75
 Appendix D
STATISTICAL SUMMARY

Between-Subjects Factors

Time 24:00 15

Control 100% t.extract 3

50% t.extract 3

75% t. extract 3

Clindamycin 3

Distilled water 3

Descriptive Statistics
Dependent Variable: Zone of Inhibition
Time Control Mean Std. Deviation N

24:00 100% t.extract 22.4667 4.86473 3

50% t.extract .0000 .00000 3

75% t. extract .0000 .00000 3

Clindamycin 32.6300 .74706 3

Distilled water .0000 .00000 3

Total 11.0193 14.48020 15

Total 100% t.extract 22.4667 4.86473 3

50% t.extract .0000 .00000 3

75% t. extract .0000 .00000 3

Clindamycin 32.6300 .74706 3

Distilled water .0000 .00000 3

Total 11.0193 14.48020 15

76
 Levene's Test of Equality of Error Variances a,b

Levene df1 df2 Sig.

Statistic

Zone_of_inhibition Based on Mean 6.535 4 10 .007

Based on Median 2.950 4 10 .075

Based on Median and with 2.950 4 2.130 .257

adjusted df

Based on trimmed mean 6.251 4 10 .009

Tests the null hypothesis that the error variance of the dependent variable is equal across groups.

a. Dependent variable: Zone of Inhibition

b. Design: Intercept + Time + Control + Time * Control

Tests of Between-Subjects Effects

Dependent Variable: Zone of Inhibition
Source Type III Sum df Mean Square F Sig. Partial Eta
of Squares Squared

Corrected 2887.018 a 4 721.755 148.977 .000 .983

Model
Intercept 1821.386 1 1821.386 375.951 .000 .974
Time .000 0 . . . .000
Control 2887.018 4 721.755 148.977 .000 .983
Time * Control .000 0 . . . .000
Error 48.447 10 4.845
Total 4756.852 15
Corrected Total 2935.466 14
a. R Squared = .983 (Adjusted R Squared = .977)

77
 Zone of Inhibition
Ryan-Einot-Gabriel-Welsch Range a,b
Control N Subset
1 2 3
50% t.extract 3 .0000
75% t. 3 .0000
extract
Distilled 3 .0000
water
100% 3 22.4667
t.extract
Clindamycin 3 32.6300
Sig. 1.000 1.000 1.000
Means for groups in homogeneous subsets are displayed.
Based on observed means.
The error term is Mean Square(Error) = 4.845.
a. Critical values are not monotonic for these data.
Substitutions have been made to ensure monotonicity. Type I
error is therefore smaller.
b. Alpha = .05.

78
79
 Appendix E
PHYTOCHEMICAL ANALYSIS

Research title: Antibacterial Activity of Tinospora cordifolia against Staphylococcus aureus

School: Far Eastern University
Address: Nicanor Reyes St, Sampaloc, Manila, 1015 Metro Manila

RESULTS OF QUALITATIVE ANALYSIS OF PHYTOCHEMICALS PRESENT IN THE ETHANOLIC

EXTRACT OF Tinospora cordifolia
(Notation: (+) = Present and (-) = Absent)
__________________________________________________

Secondary metabolite Result

__________________________________________________
Alkaloids +
Flavonoids +
Glycosides
Tannins
Amino Acids
Results:
Phytochemical investigation of the ethanolic extract of Tinospora cordifolia stems led to the presence of
Flavonoid, Glycosides, Tannins, Amino Acids and Alkaloids.

*** N o t h i n g F o l l o w s***

Remarks: All of the results of the experiment done in the research laboratory is purely based on the samples
given by the costumers. The gathered data is for educational purposes only and cannot be used for advertising
or for sales promotion.

Noted by:

Aza Freya Lan A. Tubato Ramon P. Luber, DVM, MSc

Research Assistant Chairperson
Biology, Department

80
 Appendix F
BREAKDOWN OF EXPENSES
September 24, 2018

Client Name: Far Eastern University

Title: Antibacterial Activity of Tinospora cordifolia against Staphylococcus aureus
School: Far Eastern University

PROCESS DATE RATE/LABORATORY ROOM

AND GLASSWARES USAGE
Cutting of the plants September 12, 2018
500

Grinding, Soaking and Heating of the Plants September 17, 2018 Oven: 500
Filtration + Rotary Evaporator September 19, 2018 1,000
Microwave & Hotplate: 50
Sterilization of the Culture Media and Test Rotavap: 1,100
Samples Autoclave: 350
MHA: 319.01

Pouring of the Culture Media, Inoculation of September 19, 2018 BSCII: 400
Bacteria to the Culture Media and Test Samples
Incubation of the Plates September 19, 2018 Incubator : 1,250
Phytochemical Analysis September 20, 2018 200
Checking and Recording of the Results September 20, 2018 300
Rotary Evaporator Rotavap: 800
Decontamination of the Glassware’s September 10, 2018 Autoclave: 350
Filtration + Rotary Evaporator September 25, 2018 14hrs: 1500

Service charge: ₱ 2,000

Consultation Fee: ₱ 300
Bacteria 500
Total Expenses: ₱ 11,419.01

Aza Freya Lan A. Tubato Ramon P. Luber, DVM, MSc

Research Assistant Chairperson
Biology, Department

81
 CURRICULUM VITAE

NYZEL JHEUNE Z. BAYDO

Personal Information
Address 107 Francisco St, 12th avenue, Grace Park, Caloocan City
Contact No.: 09367200198 Sex: Female
Email Address: nyzeeelbyd@gmail.com
Nickname: NJ
Birthday: June 14, 2001 Birthplace: Caloocan City
Age: 17
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Hazel Z. Baydo Father’s Name: Anthony A. Baydo

Education
Divine Mercy College Foundation Inc.
(2013-2017)
With honors

Gregoria De Jesus Elementary School

(2007-2013)

82
 MAYFLOR B. BARICAUA

Personal Information
Address: Block 16 Lot 30 Ph G3 Brgy Yakal Coty of San Jose del Monte, Bulacan, Philippines
Contact No.: 09663411557 Sex: Female
Email Address: mayflor.baricaua611@yahoo.com
Nickname: Faye
Birthday: June 11, 2001 Birthplace: Solana, Cagayan
Age: 17
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s name: Marmin B. Baricaua Father’s name: Wilson C. Baricaua

Education
Village Montessori School
(2013-2017)

Francisco Homes Elementary School

(2007-2013)

83
 ALCYREN JOY D. DE GUZMAN

Personal Information
Address 377 Alonzo Village, Cacutud, Arayat, Pampanga
Contact No.: 09351418980 Sex: Female
Email Address: deguzman.alcyrenjoy@gmail.com
Nickname: AJ
Birthday: December 07,2000 Birthplace: San Fernando, Pampanga
Age: 17
Nationality: Filipino Religion: Christian
Civil Status: Single
Mother’s Name: Lucina D. De Guzman Father’s Name: Reynaldo P. De Guzman

Education
Adelle Grace Montessori School Inc.
(2013-2017)
With high honors

Anderson Elementary School

(2007-2013)
Valedictorian

84
 GISELLE JANE C. DE GUZMAN

Personal Information
Address: Purok 6 Brgy. Sapang, Jaen, Nueva Ecija
Contact No.: 09178251114 Sex: Female
Email Address: dgisellejane0514@gmail.com
Nickname: Giselle
Birthday: May 14, 2000 Birthplace: Cabanatuan City, Nueva Ecija
Age: 18 years old
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Nenita C. De Guzman Father’s Name: Gilbert N. De Guzman

Education
General De Jesus College
(2013-2017)
With honors

General De Jesus College

(2007-2013)
5th honor

85
 SLINNIE C. ENCISO

Personal Information
Address: 150 Anahaw St. Veterans village, Project 7, Quezon City
Contact No.: 09569710305 Sex: Female
Email Address: slinniechan@yahoo.com
Nickname: Slie/Sli/Slinnie Chan
Birthday: October 19, 2000 Birthplace: San Jose Del Monte, Bulacan
Age: 17
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Annie Lynn R. Co Father’s Name: Rodel D. Enciso

Education
Ernesto Rondon High School
(2013-2017)

Esteban Abada Elementary School

(2007-2013)

86
 SOFIA LUBELLE P. LANDRITO

Personal Information
Address: Blk 1 Lot 5 Tamarind Rd. Summerfield Subd, Pasig City
Contact No.: 09475342531 Sex: Female
Email Address: sofialandrito.28@gmail.com
Nickname: Sofia
Birthday: February 28, 2002 Birthplace: Mandaluyong City
Age: 16
Nationality: Filipino Religion:
Civil Status: Single
Mother’s Name: Ma. Jennifer P. Landrito Father’s Name: Romeo P. Landrito

Education
Sacred Heart Academy of Pasig
(2013-2017)

Sacred Heart Academy of Pasig

(2008-2013)

87
 ALLIAH B. LORIO

Personal Information
Address: 2514, Jose Syguia, 884, Santa Ana, Manila
Contact No.: 09455611272 Sex: Female
Email Address: lorioalliahb3@gmail.com
Nickname: Liah/Iya
Birthday: March 14, 2001 Birthplace: San Narciso, Quezon
Age: 17
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Annelyn B. Lorio Father’s Name: Rodel A. Lorio

Education
St. Joseph’s High School
(2013-2017)

San Narciso Central Elementary School

(2007-2013)

88
 JASMINE CLAIRE M. NITUDA

Personal Information
Address: 1033 Holy Spirit Ladies Dormitory Pnoval St. Sampaloc Manila
Contact No.: 09950801509 Sex: Female
Email Address: jasminenituda@gmail.com
Nickname: Jaja
Birthday: November 8, 2000 Birthplace: Quezon City
Age: 17
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Carmela M. Nituda Father’s Name: Jessiefel L. Nituda

Education
St. Paul Academy
(2014-2017)

St. Paul Academy

(2011-2014)

Santa Clara College

(2008-2010)

89
 MERNELLE MAE O. SANTOS

Personal Information
Address: 991-G New Antipolo Tondo, Manila
Contact No.: 09293377263 Sex: Female
Email Address: mernelle_mae@yahoo.com
Nickname: Nelle
Birthday: May 26, 2000 Birthplace: Manila, Philippines
Age: 18
Nationality: Filipino Religion: Roman Catholic
Civil Status: Single
Mother’s Name: Ma. Melissa O. Santos Father’s Name: Arnel O. Santos

Education
Florentino Torres High School
(2013-2017)

Lakan dula Elementary School

(2007-2013)

90