CAVITE NATIONAL HIGH SCHOOL – SENIOR HIGH SCHOOL

Chief E. Martin St., Caridad, Cavite City

Bacteriostatic Activity of Synthesized Silver Nanoparticles and Baobab Seeds

Methanolic Extract Against Escherichia coli

A Research Paper

Presented to the Faculty of

Cavite National High School - Senior High School

Cavite City

In partial fulfillment of the requirements

for Research Capstone

CUSTODIO, JAN ANDREI F.

KALUGDAN, LUIS GABRIEL S.

VILLETE, DREANNA REMZ P.

Science, Technology, Engineering and Mathematics (STEM)

STEM 12 - E

2023 - 2024

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CHAPTER 1

INTRODUCTION

Background of the Study

The City of Cavite is a coastal area, which makes Caviteños, especially those living

near the bay, more susceptible to waterborne diseases. One example of these dangerous

diseases is Escherichia coli. Escherichia coli (E. coli) is often found in the digestive tracts of

warm-blooded animals and people (World Health Organization, 2018). The majority of E.

Coli strains are not harmful. However, some strains can lead to serious foodborne illnesses.

O157:H7, one of the most dangerous strains, can make an individual sick. It can cause

vomiting, abdominal cramps, and acute bloody diarrhea. O157:H7 is the leading cause of

acute kidney failure in children. It can also induce potentially fatal symptoms such as adult

renal failure, fever, hemorrhage, disorientation, and convulsions. (Felson, 2017). A new strain

of E.coli, referred to as REPEXH02, has been discovered recently by The Center for Disease

Control and Prevention. It is linked to numerous outbreaks of foodborne illness, particularly

those connected to romaine lettuce and other leafy greens (Reiley, 2023). It is thought to have

originated in late 2015 and has caused numerous hospitalizations and cases of hemolytic

uremic syndrome. Hemolytic uremic syndrome (HUS) develops when small blood arteries

become damaged and irritated (Mayo Clinic, 2019). HUS disrupts the kidneys’ filtering

system, which can cause kidney failure.

The Baobab tree and its related species belong to the Malvaceae family and the

Adansonia genus. The baobab is known by its distinctive appearance, broad trunk, unusual

root-like branches, and huge, velvety fruit. Due to the many distinct uses of its many sections,

the tree is steeped in tradition, and locals commonly refer to it as the "tree of life”. The name

"Baobab" is derived from the Arabic “bu hobab”, which means "fruit with many seeds"

(Jackson, 2015). The Baobab tree serves as both a source of protection and raw materials for

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numerous valuable items, such as food, clothing, and medicine. Baobab fruit pulp, seeds,

leaves, flowers, roots, and bark are all edible, and scientists have looked at their potential

purposes. Numerous biological characteristics of baobab include antibacterial, anti-malarial,

antiviral, antioxidant, and anti-inflammatory activity, among others (Rahul et al., 2015).

Baobab pulp and leaf extract synthesized with methanolic extract had similar antibacterial

and inhibitory effects to different antibiotics against Enterococcus faecalis. (El Bakkali et al.,

2022). According to a multivariate analysis, the leaves significantly reduced Staphylococcus

aureus growth and had a similar effect on antibiotics.

In recent years, nanotechnology has been a modern field of science that plays a

significant role in everyday life (Tama, 2016). As Kamur, Kaur, and Sharma (2019) stated, it

is the application of science and technology to manipulate matter at atomic and molecular

scales. Over the past two decades, studies on synthesized nanomaterials and their

characterization have been an emerging field of nanotechnology due to their vast applications

in physics, chemistry, biology, and medicine (Panorulselvam et al., 2017). In medical

science, it is used as a tool in several ways, like targeted drug delivery, gene delivery,

artificial implants, cell repair, antibacterial treatment, and diagnostic techniques. These can

reduce damage to healthy cells in the body and allow for earlier disease detection (Seybon,

2019).

Nanoparticles are described as nanotechnology's fundamental building blocks. They

are of great scientific interest as they are effectively a bridge for atomic or molecular

structures (Kamur, 2019). Catalysts are substances that change the reaction rate without being

changed by the reaction itself. Reactions take place on the surface of the catalyst. The greater

the catalyst's surface area compared with the catalyst's volume, the more influential the

catalyst will be in speeding up the reaction (Anonymous, 2019). Because of their size,

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nanoparticles have a better surface area-to-volume ratio than particles of larger size, which

causes them to be more reactive to other molecules, making them an excellent catalyst.

Various studies show that silver nanoparticles display antibacterial and antimicrobial

activity; Tama (2016) stated that AGNPs are developed as potent antibacterial, antifungal,

anti-viral, and anti-inflammatory agents. It is also the most effective nanoparticle because of

its antimicrobial efficacy against bacteria, viruses, and other eukaryotic microorganisms. In

agreement with the previous statement, Iskimani, Tavakoli, and Namaei (2017) also stated

that silver nanoparticles (AgNP's) are among the most commonly utilized nanoparticles due

to its broad antibacterial activity. For many years, AgNPs have been well-known for their

antimicrobial activity against various groups of bacteria (Guzman et al., 2021). The study by

Tama (2016) showed that the silver nanoparticles exhibited antifungal action against various

fungi. Microbes cannot develop resistance against silver because they can develop against

conventional and narrow target antibiotics. Resistance to fungal infections has emerged

recently and is a significant health problem (Mudraser, 2020). Saul (2018) also agreed that

silver nanoparticles display good conductivity, chemical stability, catalytic activity, and

antimicrobial activity. As a result of their antimicrobial properties, silver nanoparticles are

being utilized in the treatment of microbes such as bacteria, fungi, and viruses.

Studies and literature show that silver nanoparticles exhibit antibacterial and

antimicrobial activity. In contrast, gold nanoparticles are widely used in biotechnology and

the biomedical field due to their large surface area and high level of conductivity (Saul,

2016). Gold has low toxicity to biological systems, whether bacteria, animals, or humans, due

to its elemental properties (Gin et al., 2008). However, the study conducted by Zhou et al.

(2022) revealed that strong antibacterial activities were observed at all AgNP concentrations

due to their inherent elemental properties, while AuNP did not exhibit antibacterial activity

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against Escherichia coli. This shows that silver nanoparticles are more effective in inhibiting

bacteria than gold nanoparticles.

According to Chin (2019), silver nanoparticles appear yellowish brown in an aqueous

medium; as the different leaf extracts were added to the aqueous silver nitrate solution, the

color of the aqueous solution changed from faint light to brown, indicating AgNP formation.

In 2019, Kamur, Kaur, & Sharma conducted a study entitled "Synthesis,

characterization, and antibacterial potential of silver nanoparticles by Morus nigra leaf

extract." Silver nanoparticles were synthesized using Morus nigra leaf extract, and the

synthesis of silver nanoparticles was analyzed by UV Visible Spectroscopy and Scanning

Electron Microscopy. SEM analysis showed that the synthesized silver nanoparticles of

Morus nigra were 200nm and seemed to be spherical in morphology. The average particle

size was 350nm. Antibacterial activity was examined against six MTCC cultures collected

from IMTECH Chandigarh, including both gram-positive and gram-negative bacteria such as

P. aeruginosa, S. aureus, B. subtilis, E. coli, P. fluorescens, and Streptococcus mutants. Out

of these strains, the antimicrobial activity of the AgNP showed the maximum zone of

inhibition against PP. fluorescens (22 mm) and least effective against E. coli (15mm). In

concluding remarks, the silver nanoparticles synthesized using M. nigra leaf extract would be

a better antimicrobial against various bacterial species.

A study conducted by Ruta et al. (2019), entitled "Green Synthesis of Silver

Nanoparticles of Different Shapes and its Antibacterial Activity against Escherichia Coli".

Antibacterial activities of silver nanoparticles in different shapes (i.e., spherical, triangular,

and rod) were synthesized by using Red Mulberry (Morus rubra) leaf extract against

Escherichia coli in both liquid systems and on agar plates; the synthesized nanoparticles were

characterized by UV-Visible Spectroscopy, X-ray Diffraction (XRD), and Transmission

Electron Microscopy. A fluid system treated with silver nanoparticles at high concentrations

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showed delayed growth of E. coli. Hence, these findings provide a basis for measuring the

shape-dependent bacterial activity of silver nanoparticles.

With the use of nanotechnology, which is a growing trend in the field of medicine and

science, the researchers used Baobab seeds to synthesize silver nanoparticles, test if there are

any AgNP's formed by determining the color change, and test its bacteriostatic activity

against Escherichia coli at varying concentrations.

Statement of the Problem

This study aims to determine the bacteriostatic activity of synthesized silver

nanoparticles and Baobab (Adansonia digitata) leaves methanolic extract against Escherichia

coli. Specifically, it seeks to answer the following questions:

1. Is it possible to obtain a methanolic extract from Baobab seeds?

2. Is it possible to synthesize silver nanoparticles using Baobab seeds methanolic

extract?

3. Does the pure methanolic extract of Baobab seeds contain bacteriostatic activity

against E. coli?

4. Does synthesized silver nanoparticles using Baobab seeds methanolic extract exhibit

bacteriostatic activity against E. coli?

5. Which ratio between the pure methanolic extract of Baobab seeds and synthesized

silver nanoparticles exhibits the widest zone of inhibition against E. coli?

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Objectives of the Study

The main objective of this study is to determine the bacteriostatic activity of

synthesized silver nanoparticles and Baobab (Adansonia digitata) leaves methanolic extract

against Escherichia coli.

Specifically, it aims to:

1. Obtain methanolic extract from baobab seeds

2. Synthesize silver nanoparticles using Baobab seeds methanolic extract

3. Determine if the pure methanolic extract of Baobab seeds exhibits bacteriostatic

activity against E. coli.

4. Determine which ratio of pure methanolic Baobab extract and synthesized silver

nanoparticles exhibit the most significant bacteriostatic activity against E. coli.

Significance of the Study

This study will benefit a vast population of various humans, like workers,

students, and even the unemployed. Specifically, this study aims to benefit the

following populace:

Consumers. The results of this study can be the basis for treatments against

diseases caused by E. coli. Treatments will be more affordable with natural medicine

from Baobab seeds.

Business Sector. The study can be used to propagate business among eligible companies

to gain money through sales.

Environment. This study will aid the environment as it will primarily utilize Baobab

seeds that are considered waste to the environment.

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Future Researchers. This study can be used as a reference by future researchers for

further development. It can also serve as an educational resource for future researchers seeking

to explore and conduct additional investigations into the advantages of metal nanoparticles.

This study can also be a reference for exploring alternative medicinal applications from plants

and their bacteriostatic effects on bacterial strains.

Scope and Limitations

This study only focuses on synthesizing and producing silver nanoparticles (AgNP’s)

using Baobab seeds at varying proportions against Escherichia coli. The Baobab seeds will

be collected in local areas of Cavite City and will be extracted and then subjected to Rotary

Vacuum Evaporation to obtain the extract. The synthesis of AgNP’s from Baobab seeds will

happen at Cavite National High School – Senior High School. The formation of AgNP’s will

be determined by the color change of the extract from colorless to brown. The bacteriostatic

activity of the extracts against Escherichia coli will be tested at Adamson University’s

Chemical Laboratory using the Disc Diffusion Method.

Only the said materials, concentrations, and processes will be conducted. Therefore,

determining the effect of the synthesized silver nanoparticles as medicine on humans and

other subjects and testing other plant extracts, other metal nanoparticles, and other cultured

bacteria are beyond the scope of the study.

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Definition of Terms

For the reader’s clear understanding of the present study, the following terms were

defined technically and operationally:

Bacteriostatic. An agent that prevents the growth of bacteria. It is the measured or

determined characteristic in this study.

Baobab. A local plant that has therapeutic properties. In this study, it is the plant that will be

used in synthesizing silver nanoparticles.

Escherichia coli. A gram negative bacteria that is tested in this study against the treatment

(Silver nanoparticles and Baobab leaves extract). It is one of the main causes of serious

diseases.

Green synthesis. In this study, it is the process of producing nanoparticles using Baobab

seeds.

Methanolic extraction. An extraction method that is used in this study for the extraction of a

liquid substance from the Baobab seeds. In methanolic extraction, the sample material is

soaked in methanol which allows the solvent to dissolve and extract the desired compounds

from that sample.

Silver nanoparticles. Tiny particles of silver that is used as a treatment against E. coli. This

study observes this treatment.

Zone of inhibition. It is an area in the disk wherein the growth of the bacteria is inhibited. In

this study, the diameter of the zone of inhibition will determine the strength of the

bacteriostatic activity of each setup.

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