The Potential of Guava (Psidium guajava) leaf extract as an Additive

Raw Material in making Anti-fungal Bio-Based Polymer

A reasesrch project presented to the faculty of Bicol University

College of Chemical Engineering in partial fulfillment
in Chemistry for Engineers

Proponents:

Group 6 (Team Jacobi)

Juan Raymond B. Biron

Margielyn Nicole B. Briñes

Jonilyn B. Cirujales

Samuel Omar S. Gerio

Ernesto D. Gonzales V

Carl Ephraim M. Mina

Gael Laurrenz L. Nocidal

Submitted To:

Engr. Jeremiah Burac

December 2019
 ABSTRACT

This study aimed to determine the potential of Guava (Psidium guajava) leaves

decoction as additive in making anti-fungal bioplastic. It likewise determined the

effectiveness of the anti-fungal properties of the bioplastic and its other properties in terms of

tensile strength, solubility in organic and inorganic solvents, and biodegradability.

Three trials, one treatment, and three replicates were employed in the conduct of the

study, commercial plastic served as the positive control.

Results of the study revealed that there is a significant difference on the tensile

strength between set up 1 and commercial plastic, which signifies that the commercial plastic

is not comparable with the antimicrobial bioplastic in set up 1 in terms of tensile strength.

Moreover, there is a significant difference of the Anti-fungal bioplastic in organic solvent and

inorganic solvent which is water. Results showed that the bioplastic dissolved better when

subjected into inorganic solvent. Results showed that after two weeks, there is a change in the

mass of the Anti-fungal bioplastic by 1.11% when the biodegradability is tested. Based from

the statistical analysis, Guava leaves decoction can become a very good source of anti-fungal

property in an anti-fungal bioplastic.

It is therefore recommended that the correct ratio and proportion or amount of the

components of the bioplastic be further investigated for a more durable result. Further studies

must have a laboratory analysis of their test subjects for a more authentic, precise, and

accurate result.

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 Table of Contents

TITLE PAGE

Abstract 2

Acknowledgement 4

Background of the Study 5

Statement of the Problem 7

Significance or Purpose of the Study 7

Objectives 7

Significance of the Study 8

Definition of Terms 9

Scope and Limitations 10

Review of Related Literature and Related Studies 10

Related Literature 10

Related Studies 12

Materials and Methods 14

Research Design 14

Materials and Equipment 15

Treatment or Procedures 15

Results and Findings 23

Conclusions and Recommendations 34

References and Resources 35

Apendices 36

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 ACKNOWLEDGEMENT

First and foremost, we would like to thank our research coach, Engr. Jeremiah Burac.

The completion of our project would never have been accomplished if it was not for our

coaches’ guidance and involvement in assisting us through the times we had inquiries about

certain issues and problems. We would like to thank you for your support and understanding.

We would also like to thank the Chemical Engineering Laboratory (Department of

Engineering) for helping us during the pre-experimentation of the study.

Also, our deep gratitude towards Mr. Joel Illagan, the Chemical Technician of the

campus laboratory for lending us equipments and instruments to use during the course of our

study’s experiment. Consequently, we would to reach our gratitudes to the whole Chemical

Engineering Faculty for helping us with our requests during times of need.

To our parents, for their moral and financial support during the course of the study.

None of this could have happened without them.

Most importantly, we would like to thank the Lord for giving us great opportunities

and blessings. Without Him, all would have been for nothing.

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 CHAPTER 1

INTRODUCTION

1.1 Background of the Study

In the Philippines, exposure of some foods and using reusable synthetic plastics as

wrapper or container of different products are common. Example of these products which are

involved in this kind of set-up are those native kakanin and many other commercially sold

food that are found amongst streets and sidewalks. It is considered as one of the most

continued business in the country since it is affordable. This is why plastic usage is inevitable

for businesses (Phys Org, 2019).

Synthetic Plastics

Bioplastics are biodegradable materials that come from renewable sources

and can be used to reduce the problem regarding petroleum-based plastic wastes

contaminating the environment.

The world has produced over nine billion tons of plastic since the 1950s. 165

million tons were attributed to the ocean, and almost 9 million more tons were entering the

ocean each year.

Traditional synthetic plastics made from petroleum. Bioplastics are said to be

the solution to the crisis regarding occurrences of petroleum-based plastics.

More than eighteen trillion pounds of plastic have been produced to and

eighteen billion pounds of plastic flows into the ocean every year and several solutions and

plans were made . As research progresses and research concerning plastic wastes, consumers

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and manufacturers have planned to come up of a new solution as an alternative for plastics,

and then bioplastics have emerged as a potential alternative.

On 1500 BCE, MesoAmerican cultures ( Olmec, Maya, Aztecs) use natural

latex and rubber to make balls, containers and make their clothes waterproof and in 1862,

Alexander Parkes creates Parkesine the first man-made plastic made from cellulose.

It is possible to produce bioplastic out of some fruit leaves processed to

become more pathogen resistant with antiviral, antifungal, antibacterial properties.

Nafisa Jabeen (December 2015

The world almost revolve around plastics. It is widely use specially in markets and

most plastic are used in its bounds. Some of those are food packages and is a vital part of the

subject of food technology that is involved with protection and preservation of all types of

foods. Due to economical abundance, petrochemical plastics have been largely used as

packaging material due to their desirable properties of good barrier properties towards O2,

aroma compounds, tensile strength and tear strength. Meanwhile, they have many

disadvantages like very low water vapour transmission rate and the major disadvantage is that

they are non-biodegradable and result in environmental pollution and there are also risk of

producing different forms of micro-organisms that are harmful to human health. Keeping in

view the waste disposal problem of petroleum, newer cocept of use of bioplastic came into

existence. Bioplastics of renewable origin are compostable or degradable by the enzymatic

action of micro-organisms. Generally biodegradable polymers get hydrolysed into CO2, CH4,

inorganic compounds or biomass. The use of bio-origin materials obtained through microbial

fermentations, starch and cellulose has led to their tremendous innovative uses in food

packaging in the last few years.

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 Plastics are produced for convenient human consumption. Given that they are

inexpensive, lightweight, strong, durable, corrosion-resistant materials, they are used to make

vast array of products that bring us benefits (Andrady & Neal, 2009). The use of plastics have

transformed everyday life whereas it brings medical and technological advances, energy

savings, etc. As a consequence, annual production from increasing usage over the last 60

years brings concern to the environment about usage and diverse disposal, accumulation of

waste landfills and in natural habitats, problems for wildlife resulting from entanglement in

plastic, the leaching of chemicals from plastic products and the potential for plastics to

transfer chemicals to wildlife and humans. People are exposed to these chemicals not only

during manufacturing, but also by using plastic packages, because some chemicals migrate

from the plastic packaging to the foods they contain (“National Report on Human Exposure

to Environmental Chemicals," 2001). Turning to adverse effects of plastic on the human

population, there is a growing body on potential health risks. A range of chemicals that are

used in the manufacturing of plastics are known to be toxic (Talsness et al., 2009).

Many chemical additives that give plastic products desirable performance properties

also have negative environmental and human health effects. There are evidences showing that

plastic food wraps are dangerous to human health. Doctors even suggests to stop using it.

Plastics degrade or decay at a certain point (Gerrie, 2015). Direct toxicity from the release of

monomer, or other chemicals during breakdown may be added to the plastic to give it the

right mechanical properties.

Moreover, mold growth in plastic is evident. If all of the necessary conditions exist,

including the existence of dirt and excessive moisture on the surface of the plastic, as well as

a porous construction in the plastic, mold may begin to grow in as little as 24 to 48 hours

(Anzalone, n.d.). Mold are microscopic fungi that live on plant or animal matter. No one

knows how many species of fungi exist, but estimates range from tens of thousands. In

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general, it produces spores that are air, water, and vector borne (United States Department of

Agriculture, 2013). Eating mold grown in food, such as those kakanin, will likely cause some

serious diseases such as allergic reactions and respiratory problems. And, a few mold, in the

right conditions, produce "mycotoxins," poisonous substances that can make people sick

(United States Department of Agriculture, 2013).

About 420,000 people die every year worldwide because of food poisoning. At least

600 million people, or 1 in 10 worldwide, fall ill from contaminated food annually and many

of them were identified as children. Africa has the highest burden of food-borne diseases,

relative to its population. More than 91 million people are estimated to fall ill and 137,000 die

each year. Although having the second highest relative burden of food-borne diseases,

Southeast Asia suffers the most, with more than 150 million cases and 175,000 deaths a year.

60 million children under the age of five falls ill and 50,000 die and suffer from food borne

diseases (World Health Organization, 2016). Several cases of food poisoning are occuring

every year. Focusing on preventing these from happening is essential.

In connection with this, the researchers will make a bioplastic, made from accessible

biodegradable resources, to help lessen the risk of food and health contamination. To be more

specific, the researchers will aim to create a bioplastic with an anti-fungal property to inhibit

mold growth. The biopolastic will make use of Guava (Psidium guajava) leaves that is said to

have the said property. With this prospect, the researchers aim to solve problems not only in

the Philippines but in a more global context as well: addressing environmental and health

problems.

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1.2 Statement of the Problem

1. How effective is Guava (Psidium guajava) leaves decoction as an additive in making

Anti-fungal Bioplastic?

2. How effective is Anti-fungal Bioplastic in terms of tensile strength, biodegradability,

and solubility.

1.3 Goals and Objectives of the Study

Objectives

It specifically aims to:

1. To determine the potential of Guava leaves decoction as a material for the production

of anti-fungal bioplastic

2. To determine the effectiveness of the anti-fungal bioplastic in terms of:

a. tensile strength

b. biodegradability

c. solubility in organic and inorganic solvent

1.4 Significance of the Study

The study will be benefiting people in the community. This includes:

a) Home Owners

Food keeping and storing will be much easier so it will benefit

household owners. Since the bioplastic possesses an anti- fungal property, it

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 will help them store their products effectively and help maintain family

members’ safety. Considering the effects of the bioplastic, an increase in food

lifespan is an additional procurement of such household products.

b) Buyers

The most salient contribution of the product made and scrutinized is

that it will further prolong the freshness of perishable goods that will be

purchased and packaged using the bioplastic.

c) Vendors

Food businesses will be well benefited by the project. Since the bio-

plastic possesses an anti-fungal property, it will help them store their products

effectively and help maintain their buyers’ safety, considering the effects of

the bio-plastic it will also help the products of the different vendors to attain

additional lifespan of their products.

d) Future Researchers

Future researchers will be benefited by the study, by simply giving

concepts about bioplastic having an anti-fungal property. Future researchers

will have a more reined idea about anti-fungal bioplastic.

The study can also have a great impact on :

e) Environment

The biodegradability of the bioplastic is a helping plus factor to the

environment. Its capability of decomposing in a certain period of time will be

of help, given the fact that plastic-related waste is a global issue.

f) Economy

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 Philippine economy will be well benefited by this study, since the

bioplastic has anti-fungal property, it can prolong the lifespan of products.

Increasing the lifespan of products means more income to the entrepreneurs.

More income leads to a more progressive economy.

1.5 Expected Output

The expected output of the study is a tested antifungal bio-based polymer (bio

plastic) that help minimize the fungal growth of mold and likewise its environmental impact.

1.6 Scope and Delimitations

The potential of guava leaf extract as an additive in making anti-fungal bioplastic is a

study that deals with the use of guava leaf as a constituent in making cornstarch-based

bioplastic. Using this instead of water would add an anti-fungal property to the bioplastic.

The researchers tested the rectangular-shaped bioplastic in terms of its solubility in organic

and inorganic solvent, biodegradability, and anti-fungal property by subjecting it to loaf

breads. The observations took place within only five days, utilizing the complete randomized

design for the experimentation proper. The study does not not include a lab analysis of the

results of the tested food product used to determine if the bioplastic has an anti-fungal

property. Using the bioplastic in meat, vegetables and other food products are beyond the

reach of our study. The study also delimit the testing of its effectiveness in preventing

bacterial count and mold count in other products.

1.7 Definition of Terms

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a. Anti-fungal: It is a medication that limits or prevents the growth of yeasts and other

fungal organisms. It selectively eliminates fungal pathogens from a host with

minimal toxicity to the host.

b. Corn starch: It is sometimes referred to as corn flour, is a carbohydrate extracted

From the endosperm of corn. This white powdery substance is used for many

culinary, household, and industrial purposes. In the kitchen, corn starch is most often

used as a thickening agent for sauces, gravies, glazes, soups, casseroles, pies, and

other desserts

c. Solubility: It is a chemical property referring to the ability for a given substance, the

solute, to dissolve in a solvent; measured in terms of the maximum amount of solute

dissolved in a solvent at equilibrium

d. Biodegradability: It is the ability of organic substances and materials to be broken

down into simpler substances through the action of enzymes from microorganisms

e. Tensile strength: It is the ability of a material to withstand a pulling (tensile) force

f. Bioplastic: It represents a plastic substance that is based (wholly or in part) on

organic biomass rather than petroleum

g. Glycerin: It (also called glycerol) is a naturally occurring alcohol compound and a

component of many lipids. Glycerin may be of animal or vegetable origin

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 h. Organic Solvents: It is an organic solvents are carbon-based solvents that are

capable of dissolving or dispersing one or more other substances

i. Spring Scale: It is a weighing device that utilizes the relation between the applied

load and the deformation of a spring

CHAPTER 2

REVIEW OF LITERATURE AND STUDIES

2.1 Related Literature

Guava

Psidium guajava, or more specifically known as “apple guava”, are tropical trees, with

tough dark leaves that are opposite, simple, elliptic to ovate and 5–15 centimetres (2.0–5.9 in)

long. Its flowers are white, with five petals and numerous stamens; the fruits are 4-12 cm

long with round or oval shape depending on their species. The tree bears many-seeded

berries. In the Philippines, it is more commonly known as bayabas. The plant has an

economic value; its leaves, fruit and bark are used as alternative medicine to treat a variety of

diseases. Its leaves and fruits have antibacterial and antifungal properties (Latha et. al, 2013).

Anti-fungal

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 An anti-fungal is an agent that kills microorganisms or inhibits their growth.

(Anonymous, 2009). Anti-fungal products kill or slow the spread of microorganisms. Without

antifingal properties, fungal contamination can occur, which is one of the causes of food

spoilage that is a major issue for the food industry, leading to food waste, substantial

economic losses for manufacturers and consumers, and a negative impact on brand names.

Among causes, fungal contamination can be encountered at various stages of the food chain

(e.g., post-harvest, during processing or storage). Fungal development leads to food sensory

defects varying from visual deterioration to noticeable odor, flavor, or texture changes but

can also have negative health impacts via mycotoxin production by some mold. In order to

avoid microbial spoilage and thus extend product shelf life, different treatments—including

fungicides and chemical preservatives—are used. In parallel, public authorities encourage the

food industry to limit the use of these chemical compounds and develop natural methods for

food preservation. This is accompanied by a strong societal demand for ‘clean label’ food

products, as consumers are looking for more natural, less severely processed and safer

products. In this context, microbial agents corresponding to bioprotective cultures,

fermentates, culture-free supernatant or purified molecules, exhibiting antifungal activities

represent a growing interest as an alternative to chemical preservation (Salas, et a, 2017).

As mentioned, anti-fungal products kill or slow the spread of microorganisms.

Microorganisms include bacteria, viruses, protozoans, and fungi such as mold and mildew.

(National Pesticide Information Center, 2010). Mold are microscopic fungi that live on plant

or animal matter. No one knows how many species of fungi exist, but estimates range from

tens of thousands to perhaps 300,000 or more. Most are filamentous (threadlike) organisms

and the production of spores is characteristic of fungi in general. These spores can be

transported by air, water, or insects. Unlike bacteria that are one-celled, mold are made of

13
many cells and can sometimes be seen with the naked eye. Under a microscope, they look

like skinny mushrooms (United States Department of Agriculture, 2013).

Furthermore, the same department states that some mold produce mycotoxins and

aflatoxin. Mycotoxins are poisonous substances produced by certain mold found primarily in

grain and nut crops, but are also known to be on celery, grape juice, apples, and other

produce. There are many of them and scientists are continually discovering new ones. The

Food and Agriculture Organization (FAO) of the United Nations estimates that 25% of the

world’s food crops are affected by mycotoxins, of which the most notorious are aflatoxins.

On the other hand, aflatoxin is a cancer-causing poison produced by certain fungi in or on

foods and feeds, especially in field corn and peanuts. They are probably the best known and

most intensively researched mycotoxins in the world. Aflatoxins have been associated with

various diseases, such as aflatoxicosis in livestock, domestic animals, and humans throughout

the world. Many countries try to limit exposure to aflatoxin by regulating and monitoring its

presence on commodities intended for use as food and feed.

Plastics

Plastics are produced to make the lives easier but can be a threat to the world. It has

advantages and disadvantages. Plastics are either biodegradable or synthetic plastics derived

from renewable resources. Currently, petrochemical based plastics make up 99% of the

plastics market. However, production of bio-plastics is predicted to increase between 10%

and 30% of the total plastics market by 2020. These plastic materials are used as grocery

holder, drink bottles, food containers and many more. Biodegradable plastics are better than

synthetic plastics. These are seen by many as an effective solution to the world’s problem,

because they are environmental-friendly and decomposed easily. It increased soil fertility,

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low accumulation of bulky plastic materials in the environment and reduction in the cost of

waste management. Bioplastics can be recycled to useful metabolites by microorganisms and

enzymes. (Biome Bioplastics Limited, 2014).

Bioplastic

According to Cooper (2013), bioplastics are well-known over the past 15 years. It

have become an increasingly visible group of materials and its significance is being

recognized by many markets as they more frequently appear in consumer products.

Bioplastics development has been driven by drawbacks of conventional plastics, particularly

concerning use of fossil resources, recycling, littering and pollution. The origins of the

bioplastics industry go back to the mid-nineteenth century, when the first plastics ever made

were biobased; for example, celluloid, which was originally developed to replace ivory, and

casein-based plastics. However, this was soon overtaken by the development of the

petrochemical industry, which quickly became the main source for plastic materials and

technology. Consequently, the modern interest in bioplastics derives from about 15 years ago,

starting predominantly with the development of biodegradable and compostable

thermoplastics, such as polylactic acid (PLA) and polyhydroxyalkanoates (PHA). These

biodegradable materials were mostly ‘new’ polymers, compared to conventional

petrochemical polymers such as polyethylene, as well as blends with natural products such as

cellulose and starches. However, these new polymers are not necessarily biobased and many,

particularly some aliphatic polyesters, are petrochemical-based. The interest at that time was

solely on end-of-life considerations, to reduce waste going to landfill and control litter and

marine pollution, although the emphasis is now changing back to renewable sourcing, as with

the first bioplastics from the nineteenth century.

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 These ‘new’ bioplastics were designed for packaging and other applications to be

thermoplastically processable using conventional methods such as extrusion, film blowing

and casting, lamination, sheet casting and thermoforming, injection-molding, blow-molding

and foaming, both for monolithic and multilayer systems. They can also be combined with

paperboard by lamination or extrusion coating. As with conventional polymers, specific

bioplastics are selected for particular applications, depending on the requirements for:

mechanical, thermal, optical and tactile properties; chemical, oil, fat and grease resistance;

barrier properties towards oxygen, carbon dioxide, water vapor and organoleptics; adhesion

and printability; sterilizability; and cost. Also, as with conventional polymers, there is

considerable ability to tailor-make packaging to fit specific property combinations and

optimize protection of the packaged contents, while reducing costs by minimizing materials

use and maximizing processing speeds (Copper, 2013).

Polyhydroxy alkonoates (PHAs) or Polyhydroxy butyrates (PHBs) are bioplastics

which can replace a number of traditional plastics which are currently made up of

petrochemicals. The PHAs or PHBs obtained through biological origin assures the same

commercial properties with the advantage of being completely natural biodegradable. Same

way bioplastics prepared using the fruit waste mixed with glycerin will also serve as potential

alternatives to the conventional plastic materials (Yaradoddi et. al, 2016).

Glycerin

Glycerin is a colorless, odantiorless and sweet-tasting thick liquid humectant by-

product from soap making processes. Glycerin can dissolve into water or alcohol, but not in

oils. It is a good solvent; many things can dissolve easier in glycerin than they do in water

and alcohol. Glycerin is highly known as ‘hygroscopic’ which means that it absorbs water

16
from the air. Moisturizer is another word for humectant meaning, it retains moisture to the

skin so that is why it is considered to be used in commercial soaps, lotions and creams

(Westerman, 1997). In bioplastic making, glycerin acts as plasticizer meaning it makes the

polymer chain molecules in the initial product bend and slide past each other more easily,

adding flexibility into the product (Anonymous, 2014).

Starch

Starch as one of the components of bioplastic, has a heterogeneous, semi-crystalline

granular structure and the degree of ordered structure are important effect in foods and

bioplastic formation (Warren, et al., 2016). Starch is one of the biodegradable polymers

which can replace synthetic polymers in situations requiring short-term solutions, such as

food packaging and disposable cutlery. Major starch resources include potato, corn, rice,

wheat gluten and soy. In these starch is stored in the form of semi-crystalline granules

composed of two glucose polymers, amylose and amylopectin. Other recent research has

studied a reinforcement for wheat gluten based bioplastics produced from rubber and

sawdust. The experiment, carried out by Bootklad et al., evaluated the morphology, water

absorption, mechanical, thermal and biodegradation properties of the wheat gluten and found

good dispersion and uniform embedding in the material (Bootklad,2016). In this article, a few

characterization techniques were evaluated for rubber wood sawdust (RWS), such as

morphology, water absorption, mechanical, thermal and biodegradation properties of the

wheat gluten. A good dispersion and uniform embedding of the EWS was observed within

the wheat gluten matrix. With high moisture and intermediate moisture materials, such as

gels, cakes, and breads, starch bioplastics include only a limited amount of water (less than

20% w/w) both during processing and after processing (Van Soest & Knooren, 1997). Starch

structure can be characterized by differential scanning calorimetry, 13C-nuclear magnetic

17
resonance, Xray diffraction and Fourier-transform infrared spectroscopy (FTIR). Native

potato starch was derivatized using different agents to reduce its hydrophobicity. If we add

nanocellulose into the starch material, the mechanical properties of biopolymers as well as to

maintain their inherent biodegradable capability will improve (Morán, et al., 2013). This

contribution demonstrates a new approach to form bioplastics with both improved moisture

and heat resistance and high mechanical properties. Further research on morphology and

structure is essential to understand the structure-property relationship of polymer composites

(García, et al., 2009). The development of biodegradable materials to be used by the plastic

industry has been taken increasing interest over the last few years. González-Gutierrez et al

have studied the different blends of albumen protein and starch (potato and corn), which are

used as raw materials for bioplastics exhibiting high transparency and suitable mechanical

properties. However, long-term issues such as recycling bioplastic from petroleum-based

plastics, the cost, and reliability on the soil remain controversial issues. Recycling feedstock

such as pellet and natural fibres that cannot be eaten by humans and animals are good

resources in for bioplastic in terms of waste management.

Complete Randomized Design

The research design that will be used in the study is the complete randomized design.

It is an experimental design wherein subjects of the experiment is randomly assigned to

treatments. It relies of randomization to regulate the effects of the extraneous variables. When

the said experimental design is used, the proponent implies that the extraneous variables

would affect the different treatment conditions in a similar way, as to attribute any significant

difference that will be garnered by the experiment to the independent variable. It is also one

of the most commonly-used experimental designs (Hewison, 1983).

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Biodegradability

As for the parameters, biodegradability is one of the tests that will be done in the

experiment. In accordance with ASTM D 6400, biodegradable plastics are only those whose

degra- dation occurs as a result of natural action of microorganisms, such as bacteria, fungi,

and algae, in a limited period of time and in absence of ecotoxic effects’. Indeed, with regard

to recycling, nowadays processes for selecting and recycling bioplastics are not yet

developed, despite what happens for conventional plastics (Ozdamar & Ates, 2018).

Additives are often present in the bioplastics, mainly to improve the mechanical

properties of the obtained material. This choice may not only cause a reduction in the

biodegradability of plastics and other serious ecotoxic effects, but it might even determine the

non-compostability of the bioplastic, making in fact, a vain every advantage achievable

through the use of a bioplastic (Gironi & Piemonte, 2011).

Tensile Strength Test

In testing the strength of the plastic that will be made, one of the various ways to test

the strength, or mechanical behavior of a material is through the Tensile Strength Test,

usually done by placing a sample of a material that has known dimensions such as length,

within fittings that would clamp on the material. Weight is then applied to one end of the

material while the other end is fixed. The amount of weight will be increased as the measure

of the change in dimensions of the material is recorded, until such time that the material

breaks (Petroski, 1996).

Spring Scale

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 A spring scale will be used to test the tensile strength. It is a weighing device that is

consisted of a spring that is situated at one end with a hook in order for an object to be

attached. It uses applied load the deformation of a spring in order to measure the weight of an

object. It is also called spring balances, or in other cases, newton meter. It is a device used for

small-scale tensile strength tests

Hooke’s Law

The utilization of a spring scale is supported by Hooke’s Law, first used in 1676 by

British physicist Robert Hooke. It is a law of physics which states that the force needed to

extend or compress a spring by a distance is directly proportional to the scale of its distance,

mathematically stated by the formula , where k is a constant value, characterized as its

stiffness, and x is the distance of the spring (Petroski, 1996). .

2. 2 Related Studies

Anti-fungal Potency of Psidium guajava

A study conducted by Amit Pandey and Shweta from CSJM University in India

last 2011 is about the anti-fungal properties of guava leaves and fruits against various

pathogens the anti-fungal activities from fruits and leaves of guava showed lower activity in

hot water compared to methanolic, ethanolic, and ethyl acetate extracts used. But the fruits

show to have more effective anti-fungal property than its leaves (Pandey & Shweta, 2011).

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 Similarly, a study conducted by Amit Pandey and Shweta from CSJM University in

India last 2011 is about the anti-fungal properties of guava leaves and fruits against various

pathogens the anti-fungal activities from fruits and leaves of guava showed lower activity in

hot water compared to methanolic, ethanolic, and ethyl acetate extracts used. But the fruits

show to have more effective anti-fungal property than its leaves (Pandey & Shweta, 2011).

Another study from different universities in Nigeria was conducted by Fogbohun

Temitope Richard; Adekeye Temitope Joshua ; and Akinbosola Jibayo Philips that was

accepted last May, 2013.This study shows the effect of aqueous decoction of leaf and bark of

guava on fungi: Microsporumgypseum and Trichophytonmentagrophytes and on bacteria :

Staphylococcus aureus and Staphylococcus epidermidis. Mueller-Hinton agar was used to

grow the bacteria while Sabouraud Dextrose broth was used to grow the fungi. The result of

the test used in this study shows that guava solutions were effective against bacteria and fungi

(Joshua, Philips, & Richard, 2013).

Other Pharmacological Properties of Psidium guajava

A Study of the aqueous extract of kamias leaves and fruits showed antibacterial

activity against Gram-positive and Gram-negative bacteria. The antibacterial activity could

be associated with the presence of bioactive compounds of the flavonoids type, like luteolin

and apigenin. The results suggest further studies to isolate and identify the responsible

compounds (Zakaria, Z., et.al, 2007).

Likewise, Psidium guajava contains properties that can combat certain bacteria. to

verify this, the following strains of bacteria were treated with guava extract: five strains of

Listeria monocytogenes, four strains of Staphylococcus aureus, six strains of Escherichia coli

21
O157:H7, four strains of Salmonella enteritidis, one strain each of Vibrio parahaemolyticus

and Bacillus cereus, as well as five food spoilage bacteria: one strain each of Pseudomonas

aeroginosa, Pseudomonas putida, Alcaligenes faecalis, and two strains of Aeromonas

hydrophila. Among the given strains of bacteria, it was shown that the guava extract

exhibited higher antimicrobial activity in Gram-positive bacteria compared to Gram-negative

bacteria except for the following strains: V. parahaemolyticus, P. aeroginosa, and A.

hydrophila. None of the extracts showed antimicrobial activity against E. coli O157:H7 and

Salmonella Enteritidis. The minimum inhibitory concentration (MIC) of ethanol extracts of

guava showed the highest inhibition for L. monocytogenes JCM 7676 (0.1 mg/mL), S. aureus

JCM 2151 (0.1 mg/mL), S. aureus JCM 2179 (0.1 mg/mL), and V. parahaemolyticus IFO

12711 (0.1 mg/mL) and the lowest inhibition for Alcaligenes faecalis IFO 12669, Aeromonas

hydrophila NFRI 8282 (4.0 mg/mL), and A. hydrophila NFRI 8283 (4.0 mg/mL). These

results suggest that guava extract possesses compounds containing antibacterial properties

that can potentially be useful to control foodborne pathogens and spoilage organisms.

According to Mohammed, S (1994), The extracts of ripe, unripe and leaves of guava

(psidium guajava) were evaluated for antimicrobial activity against gram positive bacteria,

gram negative bacteria, yeast and fungi. Extracts from ripe guava leaves exhibited strong

activity against bacteria and in most cases, with activity stronger that 50 microgram

streptomycin.

Another study determined the antimicrobial potential of guava leaf extract

against two gram-negative bacteria (Escherichia coli and Salmonella enteritidis) and two

gram-positive bacteria (Staphylococcus aureus and Bacillus cereus), utilizing a well-diffusion

method. The guava was extracted in four different solvents, which were hexane, methanol,

ethanol, and water. The methanol extract had an antibacterial activity with mean zones of

inhibition of 8.27 and 12.3 mm, and the ethanol extract had a mean zone of inhibition of 6.11

22
and 11.0 mm against B. cereus and S. aureus, respectively. Its results showed that guava leaf

extract is a feasible natural antimicrobial agent. This, in turn, can further scrutinize other

pharmacological properties.

Another study was conducted with guava extract, this time compared with Carica

papaya in terms of their ethnomedicinal properties. An antimicrobial screening of the crude

ethanolic extracts showed activity against Staphylococcus aureus, Streptococcus faecalis, and

Escherichia coli. The minimum inhibitory concentration (MIC) for P. gujava on the organism

was found to be 5.00 mg/ml against S. aureus, E. coli and S. faecalis, while that of C. papaya

leaves is 10.00 mg/ml against S. aureus, E. coli and 8.00 mg/ml against S. faecalis

respectively. C. papaya ethanolic extract showed more active inhibition against S. aureus

with mean zone inhibition of 9.54 ± 0.03. P. gujava ethanolic extract has more active

inhibition against E. coli with antibacterial activity with mean zone of inhibition of

10.44±0.02 and S. faecalis with mean zone of inhibition of 6.72 ± 0.01 respectively. The

plants under analysis were exhibited as a good source of bioactive compounds, and thus it can

be an alternative source for garnering antibiotic substances, and therefore could be an salient

source of medicine for the treatment of various diseases.

Similar Pharmacological Properties of other plants

Likewise, a study was conducted about antimicrobial activities of decocted leaf of

guava in which the researchers were: Bipul Biswas, Kimberly Rogers, Fredrick McLaughlin,

Dwayne Daniels, and Anand Yadav from Fort Valley State University in Fort Valley, USA.

This study was accepted on the 13th day of September, 2013. In this study, guava leaves were

decocted in different solvents to show its effectiveness on two gram-negative bacteria

(Escherichia coli and Salmonella enteritidis) and two gram-positive bacteria (Staphylococcus

23
aureus and Bacillus Cereus). The result in this study shows that guava leaf-decoct can be a

natural antimicrobial agent (Anand, Bipul, Dwyane, Frederick, & Kimberly, 2013).

Furthermore, Elisa Friska Romasi, Adolf Parhusip, and Yuniwaty (2006) showed the

antibacterial activity of guava (Psidium guajava) leaves against pathogenic microbe.

According to Morton, last 2006, essential oil found in its leaves are dendrenearomatic,

selinen, nerolidiol, caryophyllene oxide, triterpenoids and sitosterol which are reported for

the antibacterial activity of its leaves. The guava leaves used in this research were obtained

from Muara Karang. From the entire experiment, it can be concluded that guava leaves have

antibacterial activity. The activity was influenced by PH, sugar, salt and by heating process. It

was tested to the four kinds of microbes: Escherichia coli, Staphylococcus aureus, Bacillus

cereus, and Penicillium using agar diffusion. Moreover, the antibacterial activity was strong

enough to inhibit B. cereus spores .This indicate that guava leaves have potential natural

antibacterial compound.

A study conducted by Wibunaranawong et al. was focused on the addition of

carvacroll to a PBS matrix as an antimicrobial agent for the preparation of food packaging.

The prepared materials showed clear zones of inhibition of bacterial growth at 4 and 10 wt%

of carvacroll; however no release studies were conducted.

Bakhshi et al. functionalized intermediate tertiary amine soybean-oil-based polyols

with ammonium salts by using either methyl iodine or benzyl chloride as alkylating agents

instead of using an acid. These bio-based polyols containing quarternary ammonium salts

were incorporated into polyurethanes using different diisocyanate monomers to obtain

biocompatible and bactericidal coatings.

Jie et al. used the extract from Scutellaria root, a herb traditionally employed in

Chinese medicine, to achieve a dual effect of dyeing and antimicrobial activity in PBS

24
matrices. Natural pigments were mixed with PBS to achieve dyed films. The materials were

tested against S. aureus bacteria, achieving antimicrobial properties at the highest pigment

load.

Synthesis of Bioplastic

Study conducted on bioplastic synthesis of chitosan and yellow pumpkin starch with

castor oil as plasticizer have been successfully synthesized by polblending which have been

printed in thi films. The resulting fim has a biodegradable characteristic for 20 days

incubation. (Hasa, et al., 2017).

Biodegradable waste was also utilized in making a bio plastic. A research study

utilized fruit waste specifically banana peels in making a bio plastic. The polymer produced

using the banana peel blended with the glycerol helped in the formation of plastic having the

characteristic features of pliability, user friendliness and strength. Other tests like solubility

and swelling studies were conducted to ensure commercial properties of these bioplastic

materials, characterization of synthesized product was carried out by FTIR and XRD

analysis, confirms the polymer is bioplastic (Yaradoddi et. al, 2016).

The use of the conventional petroleum-based plastics may have occurrence and

inhibition of contaminants, causing infection and contamination when used in food

packaging. This study investigated the antibacterial property of albumin, soy, and whey

bioplastics with the use of three common plasticizers, which are water, glycerol, and natural

rubber latex. The result is that the Glycerol infused Albumin and Glycerol infused Whey

have the most effective antibacterial activity as no bacterial growth has occurred on the

plastics after 24 hours of inoculation.

25
 Transgenically-produced amylose-only (AO) starch was used to manufacture

bioplastic prototypes. Extruded starch samples were tested for crystal residues, elasticity,

glass transition temperature, mechanical properties, molecular mass and microstructure. The

AO starch granule crystallinity was both of the B- and Vh-type, while the isogenic control

starch was mainly A-type. The first of three endothermic transitions was attributed to

gelatinization at about 60°C. The second and third peaks were identified as melting of the

starch and amylose-lipid complexes, respectively. After extrusion, the AO samples displayed

Vh- and B-type crystalline structures, the B-type polymorph being the dominant one. The AO

prototypes demonstrated a 6-fold higher mechanical stress at break and 2.5-fold higher strain

at break compared to control starch. Dynamic mechanical analysis showed a significant

increase in the storage modulus for AO samples compared to the control. The data support

the use of pure starch-based bioplastics devoid of non-polysaccharide fillers. Starch-based

bioplastic prototypes fabricated from an almost amylopectin-free starch synthesized directly

in the barley grain behaved differently compared to most high-amylose systems analyzed to

date. Melting conditions for subsequent extrusion cooking was determined from initial DSC

and X-ray scattering data. These data demonstrated effects related to interaction between the

glycerol and amylose during the heating process. A subsequent thermo-molding screen

provided optimized formulations and thermal protocols for extrusion. The finally extruded

AO starch prototypes had mechanical performances characterized by elevated stress and

strain at break compared to a control 21 starch. The AO prototypes had storage moduli

comparable to control starch prototypes, as shown by DMA data. The formulation and

processing conditions influenced the characteristics of the samples produced in terms of

molar mass distribution, residual crystallinity and mechanical properties. The data support the

use of pure starch-based bioplastics devoid of non-polysaccharide fillers. (Sagnelli, D. et al,

2017)

26
Starch-Based Synthesis

As for the methods used, a related study about bioplastic was conducted. They leaf

extraction using aqueous extraction. This is the study made by Yasmeen, A. (2011). The ratio

of the solvent (water) and the material (leaf) in the blender may be 1:10, for example (1kg:10

kg). By using aqueous extraction, homogenize the leaf and the water by blending it evenly

and then filtrate it using filter paper or heat the water with the leaves until it boils.

Whereas, according to Ozdamar&Ates (2018) in their study entitled “Rethinking

Sustainability: A research on starch based bioplastic, Journal of Sustainable Construction

Materials and Technologies”, Bioplastics are sustainable and are largely biodegradable and

biocompatible. Bioplastics can be formed in these simple steps:

Biopolymers + Plasticizers + Additives = Bioplastic

It can be formed from starch, sorbitol, glycerol (glycerin) and these ingredients are

heated to just below boiling (95°C) in a hot plate and later dried. Starch is used in its raw

state or further processed by bacterial fermentation to produce biobased monomers, which are

polymerized into Bioplastics. It usually starts with water being heated and starch is added,

water then disrupts the starch molecules. When dried, the disordered polymer chains become

entangled and a neat film is formed”. “The granules swell and burst, the semi crystalline

structure is lost and the smaller amylose molecules start leaching out of the granule, forming

a network that holds water thus increasing the mixture’s viscosity. The process itself is called

gelatinization. During the cooking of the starch, it becomes a paste that increases further in

viscosity. During cooling or prolonged storage of the paste, the semi crystalline structure

partially recovers and the starch thickens, expelling water. Vinegar (acetic acid) acts as a

27
bonding agent for the gelatin formed and the glycerin and gives it homogeneity that makes

the solution mix completely.

Glycerin

Glycerin acts as a plasticizer that intersperse among and between polymer chains

disrupting the hydrogen bonding and spreading the chains apart, which not only increases

flexibility, but also water vapour and gas permeabilities. Increasing also the plasticizer

concentration results in higher solubility (Prasad,2014).

Bioplastic Component Ratio

A study by Schon&Schwartz (n.d.) Less glycerin content increased the stiffness and

tear strength. The ratio between the mass of the water and the starch is nearly 1:1.

Temperature starts off at 65°C.

Solomonides, E. (2016). The ration of the mass of at least one starch:one

plasticizer:one acid should be at least 10:3:3 ranging from 80°C – 120 °C.

Marichelvam, M.K. et.al (2019). 40 g of corn starch were extracted and with various

ratios of glycerin, starch, gelatin and citric acid were added into 100 mL of distilled water.

The mixture was then heated using a hot plate at 100°C.

The research by Özdamara and Ateşb (2018), aimed at raising curiosity in sustainable

and experimental biobased material design by enabling an ambiguous and practice based

design process from the perspective of different disciplines. Issues on sustainability,

designing and sensing the unpredictable and searching for “new” materials for a greener and

sustainable future were the main core of bioplastic production.

As a result, a starch based bioplastic materials are constituted from vinegar, starch,

glycerol, salt and water for possible use as an emerging innovative material in architecture.

28
Regarding the negative carbon footprint and long-term environmental effects of fossil-based

plastics through landfill and incineration, the search for such a material brings forth a deeper

material experience along with a further collaboration of architects and engineering

disciplines. Through the production, they figure out deeply the nature of new starch based

materials in architecture, which are eco-friendly, cheaper and more strengthful materials

compared to conventional synthesized polymers.

Similarly, a study about corn starch-based plastic was conducted by Bernadette J.

Domingo and Scott A. Morris last March 1999. This study deals in producing a more durable

corn starch plastic with different factors like the plasticizer and processing conditions. This

said corn starch plastic degrades effectively with long time exposure to water. An hour

exposure to water made the plastic ductile, less stiff and rougher.

29
 CHAPTER 3

MATERIALS AND METHODS

3.1. Research Design

The Complete Randomized Design (CRD) was employed to determine the

effectiveness of Guava (Psidium guajava) leaves Deccoction as an Additive in making Anti-

fungal Bioplastic. Three treatments with three replicates per treatment were used in the

conduct of the study. The details are as follows:

R1 01 X 05
R2 02 06
03 07
30

04
 R3
R4 08
Let the R1, R2, and R3 have 3 slices of loaf bread
Figure 1. Complete Randomized Design

Figure 1 shows that this research study had 4 set-ups. Each set-up has the same

number of bread that was randomly assigned: R1 have 3 pieces of loaf bread, R2 have 3

pieces of loaf bread, R3 have 3 pieces of loaf bread and R4 also have 3 pieces of loaf bread.

R1 was wrapped in anti-fungal bioplastic, which was the treatment. R2 was wrapped using

paper. R3 was wrapped in commercial 8 x 11 plastic; and R4 was left uncovered. Each set up

was observed first before the application of treatment, set aside and was observed for 5 days.

In order to have efficient results, the 4 set-ups was compared and observed for 5 days. After

five days, the bioplastic was tested for anti-fungal property.

3.2. Materials and Equipment

Different materials were needed in the bioplastic making process. For the bioplastic

components, the following were used: commercially bought corn starch (60 g), guava leaf

extract (60 mL), glycerin (plasticizer; 15 mL) and distilled water (90 mL).

For the other equipments, the following were used: spoon (specifically table spoon);

non-stick pan (to use as holder for incorporating the different bioplastic components); metal

pot (where the kamias leaves will be extracted); gas stove (as a source of fire in heating or

cooking); metal spatula (used for incorporating the different components). Metal and plastic

31
containers were also used to dry the completed bioplastic to achieve a flat surface; and

weighing scale for measuring the ingredients for better and accurate results.

3.3. Treatment or Procedures

Guava leaves was collected from Brgy. Matagbac Tabaco City, Albay. After the

collection, the kamias leaves (about ½ cup) were be washed thoroughly, and were be placed

in a metal pot with tap water (half of the metal pot) to boil in moderate heat. After the liquid

is reduced to half, it is then cooled to room temperature. The leaves were then seperated with

the liquid. About 60 mL of the leaf extract will be incorporated to the bioplastic.

3.4. Manufacturing Process

In making the bioplastic, the ingredients and equipments were prepared. The pan

was heated to moderate heat. After heating, the ingredients− 50 mL of kamias leaf extract, 50

g of corn starch, 15 mL of glycerin, 15 mL vinegar and 100 mL distilled water – were placed

into the pan, respectively. The solution was heated with continuous stirring until the texture is

thick, which is the desired texture of the bioplastic mixture. The mixture was placed in square

metal containers for sun drying. The sun drying process was observed for 2 to 3 days with

relative monitoring until it solidifies.

3.5 Steps in Making Anti-Fungal Bioplastic

32
 A B

D C

A. Preparing the Bioplastic

The materials needed for the bioplastic-making process are shown: Pictures A,B, and

C shows the process of boiling the guava leaves extract; and Picture D shows the different

materials needed in order to make the Anti-fungal bioplastic.

B. Making the Corn starch-based Anti-fungal Bioplastic

The different components needed for the bioplastic-making process are incorporated in a

metal pot, as shown, to make the Anti-fungal Bioplastic. The components are measured to

make the results as desired by the team.

33
 C. Drying the Bioplastic

After drying the mixture for two to three days in a square metal tray, the result is as

shown. It is now plastic-like in its appearance. The bioplastic is brown in color due to the

guava leaves decocted color.

3.6. Schematic Diagram of Making Anti-Fungal Bioplastic

Bioplastic Production

Gathering of materials

Mixing of components

Boiling and cooking of corn starch solution

Drying of Bioplastic

34
 Figure 2. Process Flowchart for Bioplastic Synthesis

Figure 2 shows the outline of the process in making the anti-fungal bioplastic. First,

gather and organize all the ingredients and equipments and materials and prepare for the set-

up. Next, heat the frying pan (medium to high heat or 80°C-120°C). After heating, the

following ingredients were placed inside a metal pot- 50 ml of the extract, 100mL distilled

water, 50 g of corn starch, and 15mL of glycerin and 15 mL vinegar (acetic acid) . Heat the

solution and stir it constantly until it boils. If the consistency of the result is as desired, sun

dry for 2 to 3 days and monitor until it solidifies.

3.7. Experimental Set up

Anti-fungal Property Test of Bioplastic using Loaf Bread

Set up Samples Treatment

Set up 1 3 pieces of bread Wrapped using anti-fungal

bioplastic

Set up 2 3 pieces of bread Wrapped using synthetic plastic

Set up 3 3 pieces of bread Wrapped using newspaper

Set up 4 3 pieces of bread Left uncovered

Table 1. Set-up for Anti-fungal Analysis

35
 Table 1 shows four (4) different set ups with different treatments and the same

number of samples. Set up 1 has 3 pieces of bread and was wrapped using anti-fungal

bioplastic. Set up 2 has 3 pieces of bread and was wrapped using commercial plastic. Set up 3

has 3 pieces of bread and was wrapped using newspaper. Lastly, set up 4 also had 3 slices of

bread and was left uncovered.

Testing Method for Other Bioplastic Parameters

Tensile Strength

By using a spring scale, the bioplastic with a measurement of 10 in x 7 in was

compared to commercial 8 in x 11 in plastic to test for its tensile strength property. The

maximum force the bioplastic can take was determined using a spring scale.

Biodegradability Test in Soil

The bioplastic was buried inside a metal tray for two (2) weeks. The set up was left

untouched and was left inside an isolated cabinet. After the time limit, the samples were

weighed in terms of grams. The result was compared to its original weight before subjecting

it as a sample in the set up.

36
 Solubility in Organic and Inorganic Solvent

Each bioplastic was submerged in 500 mL water (inorganic solvent) and 500

mL denatured alcohol (organic solvent) for 2 weeks. The set up was kept air-tight inside an

isolated cabinet for two weeks. After that, the sample was weighed, cloth-dried first, and then

compared to its original weight before undergoing the experiment.

Schematic Diagram of the Property Tests of the Anti-fungal Bioplastic

Tensile strength

Anti-fungal property

Guava leaves decoction

Rate of decomposition

Solubility in organic and

inorganic solvent

Different kinds of variables Trials Replicates

Independent Dependent Controlled

-Tensile -Sliced bread

strength -Pure glycerine
Guava three three
-Anti-fungal -Equipments used
leaves
property -Volume of glycerine
decoction
-Rate of -Volume of guava leaves
decomposition decoction

37
 -Solubility in -Amount of cornstarch
organic and
inorganic
solvent

Table 2. Variable Distinction for the Experimentation

Table 2 shows that the decocted guava leaves directly affects: tensile strength, anti-

fungal property, rate of decomposition and the solubility (in organic and inorganic solvent) of

the bioplastic. Things that were controlled in the study were: sample number per set up, brand

of the samples, kind of corn starch, nature of the glycerin, equipments used, volume of

glycerine, volume of guava leaves extract, and amount of corn starch. Considering the

accuracy of the result of our study, we have 3 replicates and 3 trials. Having 3 trials and 3

replicates or three repetition of the study will give us concrete and efficient results.

Schematic Diagram

A. Collecting and Gathering of

the Materials

B. Making the Corn starch-based

Bioplastic

C. Testing the Corn starch-based

Bioplastic
Solubility in Organic and Inorganic
Anti-fungal Property

Tensile Strength

Biodegradability

Solvent

38
 Figure 3. Schematic Diagram for Bioplastic Synthesis

The Schematic Diagram shows the summary of the Materials and methods. The first

half was the making of the Anti-fungal bioplastic. Afterwards, the testing of the Cornstarch-

based bioplastic was conducted. The tests were to assess the bioplastic in terms of: Anti-

fungal Property, Tensile Strength, Biodegradability, and Solubility in Organic and Inorganic

Solvent. The Data Analysis or the Results and Discussions are the last part of the research

paper regarding the Effectiveness of Guava (Psidium guajava) leaves Decoction as an

Additive in making Anti-fungal Bioplastic.

39
 CHAPTER 4

RESULTS AND FINDINGS

Anti-fungal Property Test of Bioplastic using Loaf Bread (Scaling)

The Fungal Activity Before and After subjecting bread slices

% of Fungal Activity
Set Up
Before After

1 (wrapped in anti-fungal
0 0
bioplastic)

2 (wrapped in synthetic
0 50
plastic)

3 (wrapped in newspaper) 0 100

4 (left uncovered) 0 100

Table 3. Tally Table for Fungal Activity

Table 3 shows the fungal activity of slices of bread before and after five days. Seven
respondents were asked to qualitatively rate the results through the following scale:

Legend for Scaling:

- 1; if the bread has no presence of molds

- 2; if the bread has a less visible observation of molds

40
- 3; if the bread has a visible amount of molds

- 4; if the bread is full of molds

The picture shows the four set ups subjected to the anti-fungal bioplastic, commercial

synthetic plastic, newspaper, and none at all, respectively.

41
 The 3 slices of loaf bread in set up 1 showed no fungal activity. There was completely

no visual presence of molds observed.

The bread slices in set up 2 exhibited a change in the smell and appearance. There is a

less scale of fungal activity after 5 days of observation. There is a foul smell present in the

three loaf breads after the experiment.

42
 The 3 slices of loaf breads in set up 3 showed a more prominent fungal activity, as

well as a more pungent odor.

The sliced breads in set up 4 also show a large amount of fungal activity. There was

also a foul smell in the bread, and compared to the other set-ups, set up 4 had the most

prominent amount of fungal activity.

Results show that set up 1, which was wrapped in anti-fungal bioplastic, was rated 1

for microbial activity since there was completely no visual presence of of molds. Set up 2,

wrapped in plastic, was rated 2 for there were quite visible molds in the bread.However, set

up 3 and 4, which were wrapped in newspaper and was left uncovered, respectively, were

rated 3 and 4: the visual presence of molds on the bread were very prominent. Ultimately, it

43
was garnered that the bioplastic made with Guava extract showed inhibition of mold growth

in the bread.

Testing for the Other Properties of Biodegradable Plastic

Tensile Strength

1200

1000

800

600
Grams

400

200

0
Grams
Antimicrobial Bioplastic 100
Synthetic Plastic 1000

Figure 4. Comparison: Tensile Strength of Anti-fungal Bioplastic and Synthetic Plastic

44
 Figure 4 shows the tensile strength of both the Anti-fungal Bioplastic and Synthetic or

Commercial Plastic. The result shows that there was a great difference between the two tested

subjects in terms of tensile strength. The strength of the Synthetic Plastic is much greater

compared to the tensile strength of the Anti-fungal Bioplastic. The 8 x 11 synthetic plastic

held utmost 1000 grams compared to the 100 grams capacity of the Anti-fungal bioplastic

when using the Spring Scale.

Solubility in Inorganic and Organic Solvent

Original Mass Mass Loss Mass Retained

50 grams 0 grams 50 grams

Table 4. Solubility of Anti-fungal Bioplastic in Inorganic Solvent (Water) tested within 2

weeks

Table 4 and Figure 5 shows the solubility of anti-fungal bioplastic in inorganic

solvent which is water. Result shows that there is no change in its mass before and after

soaking it to inorganic solvent. Before soaking it into water, it has a mass of 50 grams and

Solubility of Anti-fungal Bioplastic in

Inorganic Solvent (Water) tested within 2
weeks
0%

Mass Loss
Mass retained
45
100%
after 2 weeks, it also has a mass of 50 grams. In conclusion the anti-fungal bioplastic is not

soluble to water.

Original Mass Mass Loss Mass Retained

50 grams 25 grams 25 grams

Table 5. Solubility of Anti-fungal Bioplastic in Organic Solvent (Denatured Alcohol)

tested within 2 weeks

Pie Chart showing the Solubility of Anti-

fungal Bioplastic in Organic Solvent
(Denatured Alcohol) tested within 2 weeks

Mass Loss
50% 50%
Mass retained

Figure 6. Pie Chart Representation of Solubility Test (Denatured Alcohol)

46
 Table 5 and Figure 6 shows the solubility of anti-fungal bioplastic in organic solvent

which is denatured alcohol. Result shows that there is a change in the mass of antimicrobial

bioplastic after soaking it into denatured alcohol. The original mass of bioplastic was 50

grams, after soaking it into denatured alcohol there was a decreased of 25 grams from its

original mass. 50% from its original mass was dissolve and 50% was retained. In conclusion

the antimicrobial bioplastic was about 50% soluble to denatured alcohol tested within 2

weeks.

Solubility of Anti-fungal Bioplastic tested within 2 weeks in Inorganic and Organic

Figure 6

60%

50%
Percentage

40%

30%

20%

10%

0%
Percentage
Inorganic Solvent(Water) 0%
Organic Solvent (Denatured
Figure 7 50%
Alcohol)

Solvent

47
 Figure 7 shows the solubility of anti-fungal bioplastic in organic and inorganic

solvent. Result shows that there was a great difference between the solubility of anti-fungal

bioplastic between inorganic and inorganic solvent. Anti-fungal bioplastic was not soluble to

inorganic solvent (water) but it was soluble to organic solvent (denatured alcohol) for about

50%. In conclusion, the anti-fungal bioplastic was soluble to organic solvent and is not

soluble to inorganic solvent that was tested within 2 weeks.

Biodegradability

Biodegradability of Anti-fungal Bioplastic tested within Two Weeks

Original Mass Mass Loss Mass Retained

50 grams 5 grams 45 grams

Table 6

Pie Chart showing the Biodegradability of Anti-

fungal Bioplastic tested within 2 weeks
1.11%

Mass Loss
Mass retained
98.89%

48
 Table 6 and Figure 8 shows that there is a change in the mass of anti-fungal bioplastic

after being buried for 2 weeks. Before burying, the anti-fungal bioplastic has a mass of 50

grams and after, result shows that there was a decrease of 5 grams to its original mass. Within

two weeks, 1.11% was degraded from its original mass and 98.89% was retained.

The Fungal Activity Before and After subjecting the slices of bread
Figure 8

D D2
Set-ups % of Microbial Activity

Before After

1 (wrapped in

anti-fungal 0 0 0 0

bioplastic)

(wrappedin 0 50 -50 2 500

synthetic plastic)

(wrapped in 0 100 -100 10 000

newspaper)

49
4 0 100 -100 10 000

(left uncovered)

Total: -250 22 500

(D and D2)

The results of the anti-fungal tests of bioplastic in terms of the presence of molds

showed that it really possesses anti-fungal property. This result corroborate with the study on

the antimicrobial activities of leaf extracts of guava in which the researchers were: Bipul

Biswas, Kimberly Rogers, Fredrick McLaughlin, Dwayne Daniels, and AnandYadav from

Fort Valley State University in Fort Valley, USA that was accepted on the 13th day of

September, 2013. In this study, guava leaves were extracted in different solvents to show its

effectiveness on two gram-negative bacteria (Escherichia coli and Salmonella enteritidis) and

two gram-positive bacteria (Staphylococcus aureus and Bacillus Cereus). Also, the study of

which concerns the testing of the leaves and bark extract of guava states that it is effective

against bacteria and fungi.

The results also corroborate with the study conducted by Fogbohun Temitope

Richard; Adekeye Temitope Joshua ; and Akinbosola Jibayo Philips that was accepted last

May, 2013.The study shows the effect of aqueous extract of leaf and bark of guava on fungi:

Microsporumgypseum and Trichophytonmentagrophytes and on bacteria : Staphylococcus

aureus and Staphylococcus epidermidis. Mueller-Hinton agar was used to grow the bacteria

while Sabouraud Dextrose broth was used to grow the fungi. The result of the test used in this

study shows that guava solutions were effective against bacteria and fungi (Joshua, Philips, &

Richard, 2013).

50
 CONCLUSIONS AND RECOMMENDATIONS

From the sufficient data garnered by the creation and experimentation of bioplastic

made with guava extract, it was found out that the bioplastic inhibited mold growth, and thus

exhibit anti-fungal properties. In hindsight, analysis of the protoype in terms of plastic

production parameters, it did not yield very favorable results. A salient case is that of the

comparison of the prototype's tensile strength with that of commercial plastic.

In order to further refurbish the study, the researchers strongly opt for finding other

ratios and components that could strengthen the bioplastic, without hindering its anti-fungal

potency. Another recomendation is the aid of laboratory institutions, to not only yield a more

quantitative scrutinization of the anti-fungal property of the bioplastic, as well as to produce a

more efficient product. The research study garnered plausive results inspite of its limitations.

51
 REFERENCE AND RESOURCES

Background of the Study:

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grow-on-plastic

2. (n.d). What Are Bioplastics?. Retrieved from.

https://www.activesustainability.com/environment/what-are-bioplastics/
3. Barrett, A. (2018). The History and Most Important Innovations of Bioplastics.
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4. Barrett, A. (2019). Technology Uses Banana Leaves as a Biodegradable Alternative to
Single-use Plastic. https://bioplasticsnews.com/2019/09/21/banana-leaves-packaging/
5. Cho, R. (2017). The Truth About Bioplastics. Retrieved from:
https://blogs.ei.columbia.edu/2017/12/13/the-truth-about-bioplastics/
6. Gibbens, S. (2018). What you need to know about plant-based plastics. Retrieved
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58
 APPENDICES
T-test:

t= D

√ ⅀D2- (⅀D)2

N (N-1)

59
 t= -250

√ 22 500 – (-250)2

4 (4-1)

t= -62.5

√ 22 500 – 15 625

12

t= -62.5

23.93567769

t= = 2.61116484 (90% level of confidence with 10% margin of error)

Data shows that the antimicrobial bioplastic is 90% effective in preventing microbial
activities with 10% margin of errors. To further strengthen our stand, here is the study
conducted by Biwas et al. which concerns about the antimicrobial activities of guava leaves
extract.

Solubility Test

A. Inorganic Solvent (Water)

60
 The Anti-fungal bioplastic is subjected into water for 2 weeks. The set up was kept
air-tight inside an isolated cabinet for two weeks. After that, we also weighed the sample
(cloth dried) and compared it to its original weight before undergoing the experiment.

B. Organic Solvent (Denatured Alcohol)

The Anti-fungal bioplastic is subjected into denatured alcohol for 2 weeks. The set up
was kept air-tight inside an isolated cabinet. After that, the sample was weighed (cloth dried)
and compared it to its original weight before undergoing the experiment.

Biodegradability/ Rate of Decomposition

61
 The Anti-fungal bioplastic made was buried inside a 1 cm metal tray for 2 weeks. The
set up was left untouched for two weeks straight inside an isolated cabinet. After the time
limit, we weighed the samples in terms of grams. The result was compared to its original
weight before subjecting it as a sample in the set up.

62