EVALUATION OF Lemna minor (COMMON DUCKWEED) MEAL AND

Glycine max (SOY BEAN) AS POTENTIAL PROTEIN ALTERNATIVE

FOR FISH MEAL IN Oreochromis niloticus (TILAPIA)

FINGERLINGS FORMULATED DIET

A Science Investigatory project of

POLOMOLOK NATIONAL HIGH SCHOOL

Octavio Village, Cannery Site, Polomolok, South Cotabato

(LIFE SCIENCE - TEAM)

By:

DELA CRUZ, ALLYZA KAYE A.

FABULARE, LEONAH JANE S.
RODEROS, JHAMAICA B.
SANDOY, ELLORVIE C.
Researchers

ARDON SABALOSA

Adviser

June 2022
RESEARCH
PROPOSAL
 TABLE OF CONTENTS

Page

TITLE PAGE…………………………………………………………………….. i

TABLE OF CONTENTS .……………………..……………………………….. iii

A. EXECUTIVE SUMMARY……..…………………………………………… v

B. INTRODUCTION

Rationale…………………………………………………………..…………… 1

Significance of the Study…………………………………………..…………... 2

Scientific Basis……………………………………….……………..……...…... 3

Theoretical Framework……………………………………………….…..…… 5

Statement of the Problem…….……………………………………….……….. 6

C. REVIEW OF RELATED LITERATURE

Related Studies……..…………………………………………………………. 7

D. METHODOLOGY

Research Design……………………………………………………………….. 17

Experimental Process Chart ..………………………………………………….. 19

Research Locale ……….……………………………………………………. 20

Data Gathering Procedures………………………………………………...….... 20

Conceptual Framework………………………………………………………..... 26

iii
E. EXPECTED OUTPUT AND POTENTIAL

IMPACT………………………………………..……………………………. 27

REFERENCES………………………………………………………….…… 30

CURICULUM VITAE……………………………………………………… 41

iv
 A. EXECUTIVE SUMMMARY

Title: Evaluation of Lemna minor (Common Duckweed) Meal and

Glycine max (Soybean) Meal as Potential Protein
Alternative for Fish Meal in Oreochromis niloticus (Tilapia)
Fingerlings Formulated Diet

Researchers: Allyza Kaye A. Dela Cruz, Leonah Jane S. Fabulare,

Jhamaica B. Roderos, and Ellorvie C. Sandoy

School: Polomolok National High School

Brgy. Cannery Site, Polomolok, South Cotabato

Fish meals serve as a foundation and source of nutrients such as protein, lipids

(oils), minerals, and vitamins. Protein, obtained from fish meals, is an essential part of

fish nourishment and due to the increased demand of commercial fish meal the price

also increases. Aside from the increased price, the production of fish meals has also

negative environmental effects. As a buttress, fish meals has been identified as impact

hotspot and is one of the parameters that contribute to the overall environmental

issues such as water eutrophication, water quality, and destruction of natural habitats

induced by aquaculture marine-based protein production.

As to mitigate the various problems entailed with commercial fish meals,

alternative protein source can be employed. Copious source of Lemna minor

(Duckweed) and Glycine max (Soybean) in the Philippines can be used as an

alternative protein source to the high cost commercial fish meal, in this way the

mitigation of cost and environmental problems can be realized.

v
 Furthermore, this study will utilize mix method Experimental type of research.

There are various tests that will be conducted inclined in the evaluation of Lemna

minor (Common Duckweed) meal and Glycine max (Soybean) meal as potential

protein alternative for fish meal in Oreochromis niloticus (Tilapia) fingerlings

formulated diet. The first test will be the evaluation of the protein concentration of

each treatment with the use of proximity test. The second test will be the assessment

which among the treatments is more potent in terms of growth effect. The third test

will be the evaluation if there are any significant relationship between the protein

content and growth effect of each treatment. Finally, evaluating if there are any

significant differences in the evaluation of the potentiality of alternative protein

source among the treatments in terms of their protein concentration and growth effect

are the fourth and last test.

vi
 B. INTRODUCTION

Rationale

Oreochromis niloticus (Tilapia) is deemed to be the second most sought

farmed fish world-wide and it was known to quadrupled its production for the past

decades considering its suitability for aquaculture, marketability, as well as stable

market prices (Wang & Lu, 2016). This species of fish is considered as one of the

most significant among others in tropical and sub-tropical aquaculture (FAO, 2012).

Considering their characteristics to reproduce and grow steadfastly during wide range

environmental conditions and tolerate handling stress (Siddik et al., 2014) they have

become an excellent choice for aquaculture. With this species significance, it is only

right that their fish feeds are both sustainable in economic and environmental aspect.

Essential nutrition among fishes is one of the crucial necessities when it comes

to aquaculture considering that it serves as the greatest factor for the growth and

nourishment of the fishes. Aquaculture is an industry of animal production that has

the highest growth rate for the past 20 years wherein the most significant activities are

predominantly grounded on fish nutrition (Dantas et al., 2017). In fact, about 40% of

the total production costs of aquaculture are set for the feeding of the fish only to

avoid future losses due to inefficiencies, especially for the intensive aquaculture

systems (Zhou et al., 2017).

Furthermore, fish meal is deemed as a feed ingredient with high quality, great

quantities of energy per unit weight, and is digestible which is fed to various animals

including mink, dogs, cats, and cattle but preferably used for fish diet. Fish meals

serve as a foundation and source of nutrients such as protein, lipids (oils), minerals,

and vitamins. Protein, obtained from fish meals, is an essential part of fish

1
nourishment and due to the increased demand of fish meal the price also increases

(Hyacinth et al., 2019).

Aside from the increased price of commercial fish meals, its production has

also negative environmental problems. Brought by the rapid expansion of aquaculture,

commercial food production systems world-wide is the critical concern in the

sustainability of the environment. The production of commercial fish meals has been

identified as impact hotspot and is one of the parameters that contribute to the overall

environmental issues induced by aquaculture marine-based protein production

(Ghamkhar & Hicks, 2020).

As to mitigate the aforementioned various problems entailed with commercial

fish meals, alternative protein source can be employed. Alternative protein source is

derived from plants, insects, or algae (Riddick & Mahboob, 2014). Copious source of

Lemna minor (Duckweeds) and Glycine max (Soybean) in the Philippines can be used

as an alternative protein source to the high cost commercial fish meal, in this way the

mitigation of cost and environmental problems can be realized.

Significance of the Study

This study is significant in testing the potentiality of Lemna minor

(Duckweed) and Glycine max (Soybean) as alternative source of protein. If proven to

be a good source of protein, these nutrients are widely available that can be used as

alternative quality source of protein. Furthermore, to produce an alternative protein

source that cannot affect the marine biodiversity. The researchers also wanted to

produce a kind of protein that is low cost, affordable, and at the same time healthy.

2
 Society. This study may help the society to understand and learn how alternative

protein source is important not only to our lives and environment but also to the basis

of aquaculture.

Bureau of Fisheries and Aquatic Resources. With this research, the Bureau

of Fisheries and Aquatic Resources will be guided and value the usefulness of Lemna

minor (Duckweed) and Glycine max (Soybean) as potential protein alternative for fish

meal. In this way, they can improve and create fish meals that are cost-mitigated and

environmental friendly.

Aquaculture Farmers. Significantly, the result of this study will directly help

the aquaculture farmers by providing them an alternative fish meal that will mitigate

their expenses and will deter negative environmental factors.

Researchers. The findings and results of this study may help future

researchers, with their similar researches and may provide new information that could

be applied on their studies. It would open new doors for much more unique and

beneficial research studies, and new ideas worthy of in-depth research and analysis.

And this could be used as basis, additional source, and reference.

Scientific Basis

Various studies have already been established similar to this specific study. One

of it is the study conducted by Harshani et al. (2020) which focuses on the evaluating

of the use of Lemna minor (Duckweed) as a formulated fish meal. This study

determined the influence of Lemna minor (Duckweed) on Oreochromis niloticus

(Tilapia) growth. A total of six treatments were used in this study wherein treatment 1

is purely commercial fish meal and treatments 2-6 are dried Lemna minor

3
(Duckweed) meal, which replaced 10%, 20%, 30%, 40%, and 50% of the fish meal.

The reported mean weight gain of the fish in each experimental diet 2, 3, 4, 5, and 6

was significantly different from that measured in control diet (p<0.05). The Specific

Growth Rate of fish which was fed with the control diet and the experimental diets

were significantly different. The findings demonstrate that the inclusion of about 30%

dried Lemna minor (Duckweed) meal in a designed diet for Oreochromis niloticus

(Tilapia) fish is highly effective and has the fastest development rate.

Another research was dedicated in evaluating the effects of substitution of fish

meal by Glycine max (Soybean) in Oreochromis niloticus (Tilapia) diet. The

experiment of Ahmad et al. (2020) was conducted in saline water to assess the effect

of replacing fish meal with a locally accessible plant protein source, Glycine max

(Soybean) meal, on Oreochromis niloticus (Tilapia) juveniles’ growth performance,

survival, and body composition. Treatment 1, 2, 3, and 4 were four types of diets in

which fish meal was partially substituted with Glycine max (Soybean) meal up to 0%,

25%, 50%, and 75%, respectively. When comparing the results of the experimental

treatments to the control diet, there were no significant differences (P>0.05) in percent

weight gain (percent), specific growth rate (SGR percent) and average daily weight

gain (g/ind/day). As a conclusion, 50% Glycine max (Soybean) meal in the diet can

replace fish meal without compromising the health and development of Oreochromis

niloticus (Tilapia).

Both alternative protein sources are proven to have contribution in the growth of

the fish, specifically the species of Oreochromis niloticus (Tilapia), it therefore

4
implies that Lemna minor (Duckweed) and Glycine max (Soybean) can be used as

formulated fish meals.

Theoretical Framework

Lemna minor (Duckweed) is known as a high-yielding plant that has been

thoroughly researched for its use as a protein source in food and feed industry

(Appenroth et al., 2017; Putra and Ritonga, 2018). In fact, harvested Lemna minor

(Duckweed) plants contain up to 16%-45% crude protein on a dry weight and may be

used without further processing as a complete feeds for fish (Zhao et al., 2014). It

therefore suggests that Lemna minor (Duckweed) can be used as an alternative protein

source for fish meal.

Furthermore, Glycine max (Soybean) can also be considered as one of the potent

substitute to fish meal considering its high protein concentration, adequate supply,

and reasonable price (Acar et al., 2013; Pervin et al., 2020; Meng et al., 2020). It can

be used as viable alternative protein sources in aqua feeds, either partially or

completely replacing fish meal (Zhou et al., 2018) with an average crude protein of

40-44%.

5
Statement of the Problem

The purpose of this study is to evaluate the potentiality of Lemna minor

(Duckweed) and Glycine max (Soybean) as potential protein alternative for fish meal.

Specifically this study aims to seek answers for the following questions:

1. What is the protein concentration in each treatment of the formulated fish

meals?

2. Which among of the treatments are more potent as an alternative protein

source in terms of their growth (weight) effect?

3. Is there any significant relationship in each treatment between their protein

concentration and growth (weight) effect?

a. 100% Commercial fish meal

b. 100% Lemna minor (Duckweed) meal

c. 100% Glycine max (Soybean) meal

d. 50% Lemna minor (Duckweed) meal and 50% Glycine max

(Soybean) meal

e. 50% Commercial fish meal and 50% Lemna minor (Duckweed) meal

f. 50% Commercial fish meal and 50% Glycine max (Soybean) meal

4. Is there any significant difference in the evaluation of the potentiality of

alternative protein source among the treatments in terms of their protein

concentration?

5. Is there any significant difference in the evaluation of the potentiality of

alternative protein source among the treatments in terms of their growth

(weight)?

6
 C. REVIEW OF RELATED LITERATURE

DUCKWEED

Lemna minor

Kingdom: Plantae

Clade: Tracheophytes

Clade: Angiosperms

Clade: Monocots

Order: Alismatales

Family: Araceae

Subfamily: Lemnoideae

Genus: Lemna

Species: minor

Lemna minor (lesser duckweed—Angiospermae, Lemnaceae) is a freshwater

plant that can be found in almost every country on the planet, primarily in lowlands

and foothill locations with stagnant or slow-flowing water. These little floating

macrophytes deemed as fastest growing Angiosperms, with a growth rate that is

nearly exponential (Ziegler et al., 2015). The body of the plant is made up of a long

base and 2 to 5 mm oval-shaped leaves that float on the water's surface and because of

its low fiber content it is easily digestible by monogastric mammals and fish (Aslam

et al., 2017). The plant is usually found in colonies of two to five leaves. Under

optimum conditions in nutrient-rich waters, a little solitary plant can reproduce every

7
three days (Kuznetsova, Politaeva, Smyatskaya, & Ivanova, 2019). Lemna minor

(Duckweed) also grows quickly, can withstand a wide range of temperature and pH

conditions, and are simple to maintain and harvest, making them a good choice for

wastewater bioremediation (Kaur & Kanwar, 2022). Such characteristics have made

the Lemna minor (Duckweed) a useful plant in various ways.

Moreover, with the various issues entailed with commercial fish meal

especially in economic and environmental aspect, it is significant to provide

alternative protein source to mitigate these problems and Lemna minor (Duckweed)

can be used as possible protein substitute for that. Lemna minor (Duckweed) is known

as a high-yielding plant that has been thoroughly researched for use as a protein

source in food and feed industry (Appenroth et al., 2017; Putra and Ritonga, 2018). In

fact, the results of a specific study demonstrated that replacing 25% of the fish meal in

the diet of Oreochromis niloticus (Tilapia) with Lemna minor (Duckweed) meal

promotes greater growth and feed utilization than feeding only fish meal, potentially

lowering the cost of feeding fish and increasing profit margins. Additionally, the

plant’s greatest potential, lies in its ability to produce large protein-rich biomass

which can be used to feed livestock, wherein the harvested Lemna minor (Duckweed)

plants contain up to 16%-45% crude protein on a dry weight and may be used without

further processing as a complete feeds for fish (Zhao et al., 2014). It, therefore,

implies that Lemna minor (Duckweed) can reduce the problem of farmland scarcity

for the production of animal feeds.

8
SOYBEAN

Glycine max

Kingdom: Plantae

Clade: Tracheophytes

Clade: Angiosperms

Clade: Eudicots

Clade: Rosids

Order: Fabales

Family: Fabaceae

Subfamily: Faboideae

Genus: Glycine

Species: max

Soy is an excellent source of protein for vegetarians, vegans, and anyone who

wants to reduce their meat intake. Soy protein products can be good alternatives for

animal products, according to the US Food and Drug Administration, since, unlike

some other legumes, soy has a "complete" protein profile (Health Jade 2018). Glycine

max (Soybean are legumes that grow all over the world and are one of the world's

most important food crops. Tofu, miso, soymilk, and tempeh are just a few of the

dishes made from them (McGuigan, 2022). It is a unique leguminous crop since its

seed is a rich source of protein, essential amino acids, oil, and metabolisable energy.

9
The two most cost-effective soybean components are oil and Glycine max (Soybean)

meal (Warrington et al. 2015).

Moreover, Glycine max (Soybean) meal is a primary protein source for animal

feed, and it is normally traded based on its protein content. The removal of the hulls at

the crushing plant can improve the protein content, hence dehulled Glycine max

(Soybean) meal should have a greater protein content than non-dehulled Glycine max

(Soybean) meal (Tangendjaja, 2015). Additionally, Glycine max (Soybean) seeds

have high protein content and a nutritionally balanced amino acid profile, they are

commonly used as a primary source of vegetable protein around the world. (Zhang,

D., Lü, H., Chu, S. et al. 2017).

Fish meal is widely employed as an animal protein source in the aquafeed

industry due to its excellent nutritional content, palatability, and enhanced

digestibility (Dossou et al., 2018a, b). However, resource scarcity and growing prices

severely limit the usage of fish meal in food fish farming (Dawood et al., 2020a; Bu et

al., 2018). In light of this, numerous efforts have been made to find sustainable

protein alternatives to replace fish meal. Because of the consistent supply and

inexpensive pricing, plant proteins are increasingly being employed in aquaculture as

a substitute for fish meal (Dawood et al., 2015; Ismail et al., 2019).

Due to its highly digestible protein and amino acid profiles, Glycine max

(Soybean) meal is the most often utilized alternative protein source to fish meal in fish

and shrimp diets (Karthik Masagounder, 2016). Glycine max (Soybean), a major

vegetable source alternative to fish meal, have been widely used in feeds for the

majority of farmed fish because of its relatively high protein quality, steady supply,

and low price (Drew et al., 2017). It is also the most widely used by-product in the

10
feed sector due to its large manufacturing and wide applicability (Opapeju et al.,

2016; Gandhi, 2019).

Glycine max (Soybean) meal, as a plant protein, is considered to be one of the

most hopeful candidates to substitute fish meal. Compared with other plant protein

sources, it has a comparatively high content of effective protein, relatively balanced

amino acid profile, steady supply, and reasonable price (Acar et al., 2013; Ye et al.,

2019; Pervin et al., 2020; Meng et al., 2020).

11
FISH MEAL

Fish meal is used as a conventional source of protein in poultry feed and is a

widely used feed for shrimp, farmed fish, and other animals around the world. It is

being trusted as the most reliable protein source due to its nutritional quality in terms

of palatability, digestibility, and excellent nutrient profiles to fulfill the dietary

requirement of aquatic species (Jannathulla et al., 2019). Particularly, its production

has been centered on a few top producers; in 2007, the top ten manufacturers

accounted for almost 80% of global production. Peru was the major producer, with

China coming in second, Chile third, and the Nordic countries of Norway, Denmark,

and Iceland coming in fourth. From nearly 33 million tons of whole fish and

trimmings, about 300 dedicated plants worldwide create about 6.3 million tons of fish

meal and 1.1 million tons of oil annually (FIN 2012).

Fish meal was a ubiquitous ingredient. In fact it is a commodity product with

millions of tons of it being traded each year on international markets and is present in

many of the products we all regularly buy. And yet at the same time it is a source of

terrible environmental damage and human suffering. Brought by the rapid expansion

of aquaculture, commercial food production systems world-wide is the critical

concern in the sustainability of the environment. The production of commercial fish

12
meals has been identified as impact hotspot and is one of the parameters that

contribute to the overall environmental issues induced by aquaculture marine-based

protein production (Ghamkhar & Hicks, 2020). Additionally, the production of

commercial fish meal has led to declined native fish stocks, high disease rate, as well

as coastal habitat damage (Henry, 2015).

Each year the UN’s Food and Agriculture Organization (FAO) produces a

long publication called the ―State of the World Fisheries and Aquaculture‖ report. The

2014 edition says that fishmeal is made from residues from fish factories (35%) and

fish that are caught for the purpose. A staggering 20% of the total fish catch is used

directly to make fish meal or fish oil. That amounts to more than 20 million tons of

wet fish from the sea.

Every responsible aquaculture industry focuses on lowering production costs,

increasing production system efficiency, and promoting environmental sustainability.

Given that fish feeds are one of the most expensive inputs in aquaculture production,

it is critical to supply fish feeds that are cost-effective, environmentally friendly, and

nutritionally adequate. Because of its nutritional quality in terms of palatability,

digestibility, and excellent nutrient profiles, fish meal has been a popular protein

source in fulfilling the dietary requirement of aquatic species (Jannathulla et al.,

2019). However, due to dwindling catch fisheries, intense competition between the

human and animal industries, global increases in energy costs, and unpredictable

year-round supply, fish meal has become increasingly limited and expensive (FAO,

2013). As a result, the widespread use of fish meal in aquaculture has not only

jeopardized the long-term viability of fisheries ecosystems, but it has also boosted fish

demand, reducing fish farmers' profit margins. (Ogello, Munguti, Sakakura, &

Hagiwara, 2014).
13
 Moreover, processed fish meal and its constituent ingredients are in greater

demand as the global aquaculture sector expands. However, the rising costs of fish

meal and fish oil will limit aquaculture's future growth. To address this issue,

scientific research and feed makers have made tremendous progress in developing

feeds that provide appropriate nutrition for animal growth especially in aquatic

species (Gasco et al., 2018). Since commercial fish meal has been expensive to

produce it is only significant to find new protein sources for fish diets which will be

vital to the aquaculture industry's economic success. One of the possible replacement

as a fish meal is the Microalgae considering that it gives an adequate source of

protein, vitamins, and essential fatty acids, particularly in tropical climes where

microalgae is abundant (Ayoola, 2013; El-Hindawy et al., 2012).

14
TILAPIA

Oreochromis niloticus

Kingdom: Animalia

Phylum: Chordata

Class: Actinopterygii

Order: Cichliformes

Family: Cichlidae

Genus: Oreochromis

Species: niloticus

Oreochromis niloticus (Tilapia) has the highest number of mass and family

based breeding programs in aquaculture, with a remarkable genetic progress realized

over the last few decades (Gjedrem et al., 2012; Komen & Trong, 2014). It is

considered as a standout amongst the most critical freshwater species for commercial

aquaculture because of its high nutritional qualities, quick development rate, and

resistance to illnesses (Nakkina, 2016). This species of fish are excellent in

aquaculture considering that little input is required and the technology is simple and

easy to adapt for small-scale fish farmers. Oreochromis niloticus (Tilapia) farming is

also profitable for investors due to its low investment cost and high feeding efficiency

in lakes and ponds where natural food is available (Romana, Eguia, & Pakingking,

2020).

15
 Globally, Oreochromis niloticus (Tilapia) and its hybrids are the most

cultivated and widely farmed fish freshwater species after carps. It is the most

important species, accounting for more than 90% of the total Oreochromis niloticus

(Tilapia) production (Fitzsimmons, 2016). Moreover, Oreochromis niloticus (Tilapia)

was the most frequently cultured species, with a 7%-10% growth in output over the

last 20 years (Fitzsimmons, 2013). They are the second largest users of commercial

fish meals after carp (6.67 and 11.03 million tons, respectively) and account for about

3% of world fish meal supply (Tacon et al., 2006). Additionally, about 75 percent of

Oreochromis niloticus (Tilapia) produced globally were fed with commercial fish

meal in 2000, and that figure will have increased to 100 percent by 2025 (Tacon &

Metian, 2015). Because Oreochromis niloticus (Tilapia) production is expected to

double from 4.3 million tons to 7.3 million tons between 2010 and 2030 (World Bank,

2013), it is necessary that we find an alternative to expensive, ecologically harmful,

and unnecessary ingredients in Oreochromis niloticus (Tilapia) fish meals.

16
 D. METHODOLOGY

This chapter is specifically intended for research-inclined information that

contains the research design, research locale, materials and instrumentation, as well as

the data gathering procedure that will be utilized by the researchers in this particular

study. This elaborates and further explains the methods that will be used in the

experiment to attain valid and relevant results for the study.

Research Design

This section covers the design on how the procedures and methods of the

study will be done. This study will utilize mix method Experimental Type of research.

Specifically, the researcher will use a True Experimental mix method Type of

research.

This study will take several processes and tests in evaluating the Lemna minor

(Duckweed) Meal and Glycine max (Soybean) Meal as Potential Protein Alternative

for Fish Meal in Oreochromis niloticus (Tilapia) Fingerlings Formulated Diet.

17
Research Design

Evaluation of Potential Protein

Alternative for Fish Meal

T1 - 100% Comercial Fish Meal

R1
T2 - 100% Lemna minor
T1 R2
(Duckweed) Meal
R3
T3 - 100% Glycine max
(Soybean) Meal
R1
T2 R2
T4 - 50% Lemna minor
R3
(Duckweeds) Meal and 50%
Glycine max (Soybean) Meal
R1
T3 R2 T5 - 50% Commercial Fish Meal
R3 and 50% Lemna minor
(Duckweeds) Meal
R1
T4 R2 T6 - 50% Commercial Fish Meal
R3 and 50% Glycine max (Soybean)

R1
Meal

T5 R1
R1 R - Replication

R1
T6 R1
R1

Figure 1 shows the chief design of the research wherein various treatments

will be tested on Oreochromis niloticus (Tilapia) fingerlings to know the potentiality

of Lemna minor (Duckweed) and Glycine max (Soybean) as an alternative protein

18
source for fish meal. There are a total of six treatments for the test and each treatment

has three replications.

Experimental Process Chart

Below is a schematic diagram that shows the different steps in the whole

procedure that will be observed while conducting this specific research.

Preparation of Collection of Drying Procedures

Materials alternative protein
sources

Proximity Test
Pulverizing Weighing

Testing the Cleaning and

potentiality of the Observation Proper Disposal
alternative protein
sources

Statistical Analysis

Figure 2 shows the various steps that will be followed by the researchers in

conducting this study. Through this, systematic process operation of the research will

be realized.

19
Research Locale

The research locale of this study will be at Polomolok National High School

situated at Octavio Village, Barangay Cannery Site, Polomolok, South Cotabato

particularly at Senior High School Research Dome.

Data Gathering Procedures

This section shows the procedures that will be followed by the researchers in

conducting the study.

Materials and Instrumentation

 Mortar and Pestle

 Microwave

 Fish Net

 Aerators

 Electronic Precision Balance

 Petri Dish

 5 kilos Lemna minor (Duckweeds)

 5 kilos Glycine max (Soybean)

Experimental Specimen

 Lemna minor (Duckweed) meal

 Glycine max (Soybean) meal

 Oreochromis niloticus (Tilapia) fingerlings

20
Expiremental Design

The study will be laid out in a Complete Randomized Design (CRD).

Treatments will be varied according to the following:

T1 – 100% Commercial fish meal

T2 - 100% Lemna minor (Duckweed) meal

T3 – 100% Glycine max (Soybean) meal

T4 - 50% Lemna minor (Duckweed) meal and 50% Glycine max (Soybean) meal

T5 - 50% Commercial fish meal and 50% Lemna minor (Duckweeds) meal

T6 - 50% Commercial fish meal and 50% Glycine max (Soybean) meal

A. Gathering of Experimental Treatments

A.1 Gathering of Lemna minor (Duckweed)

The Lemna minor (Duckweed) will be purchased in Barangay Lumakil,

Polomolok, South Cotabato. Identification and classification will be done through

internet and book research.

A.2 Gathering of Glycine max (Soybean)

The Glycine max (Soybean) will be collected in Citizen’s Village located in

Polomolok, South Cotabato, wherein, supply of Glycine max (Soybean) is available.

B. Preparation of Experimental Treatments

B.1 Preparation of Lemna minor (Duckweed)

After purchasing and harvesting the Lemna minor (Duckweed) it will be then

collected and scattered on a rice bag to be sun-dried. It will take the researchers

21
around 1–3 days to completely evaporate the moisture in the raw material, depending

on the day's weather. Afterwards, the process will be completed by microwaving the

dried Lemna minor (Duckweed) for 20- 35 minutes to thoroughly dry it.

B.2 Preparation of Glycine max (Soybean)

The preparation process for the Glycine max (Soybean) will consist the

removing of dirt and sun drying it for about 15- 25 minutes, or until the Glycine max

(Soybean) will completely dry. After that, they will be heated to be thoroughly dried

quickly, with the use of microwave and a wrap of foil. The time of heating will

depend on the moisture and texture of Glycine max (Soybean), approximately 30

minutes to 45 minutes or until it can be pulverized.

C. Pulverizing Process

C.1 Pulverizing Process of Lemna minor (Duckweed)

In pulverizing the Lemna minor (Duckweed), the researchers will prepare two

simple tools; mortar and pestle. For the Lemna minor (Duckweed) to turn into

powder, the researchers will need to thoroughly crush and grind it into fine powder.

The process will be done by gradually putting adequate amount of dried Lemna minor

(Duckweed) inside the tool. Afterward, it will be pulverized for about 15-20 minutes

or until it will be crushed entirely.

C.2 Pulverizing Process of Glycine max (Soybean)

The pulverizing of Glycine max (Soybean) will be done with certain and

systematic process. Initially, after the Glycine max (Soybean) will dry, the researchers

will utilize the tool- mortar and pestle to totally pulverize them. Initially, smaller

amount of Glycine max (Soybean) will be gradually putted inside the tool to be

22
subjected to pounding. Afterward, the Glycine max (Soybean) will be pulverized for

about 15-25 minutes.

D. Preparation for Actual In-Vivo Experimentation

The Senior High School Research Dome of Polomolok National High School

will be used for the growth trial. A pond sectioned into six parts using fish net will

serve as experimental tanks for each treatment. Tap water will be supplied in the pond

during the experimental period and aeration will be provided to maintain the adequate

oxygen level. Induced bred of Oreochromis niloticus (Tilapia) fingerlings will be

obtained from a stocking pond from Barangay Lumakil, Polomolok, South Cotabato.

Prior to the start of the experiment, the fingerlings will be acclimated and conditioned

in the pond for two days. The feeding trial will last for a month, in that span of time of

time the researchers will be able track the growth progress of Oreochromis niloticus

(Tilapia) fingerlings.

E. Feeding Procedure

The Oreochromis niloticus (Tilapia) fingerlings will be randomly distributed

at the rate of 5 fishes in each replication. The fish will be fed at satiation level with

the formulated diets on a scale of 0.3 grams three times daily at 4 hourly intervals

between 08:00 a.m. to 4:00 p.m. In order to maintain the oxygen quality, aerators will

be constantly monitored throughout the experimental period in order to prevent

overheatin

23
F. Weighing Procedure

The weighing of the experimental formulated treatments that will be fed on the

experimental specimen: Oreochromis niloticus (Tilapia) fingerlings will be realized

using the Electronic Precision Balance for more accurate scale. Similarly, the same

weighing instrument will be used in recording and attaining the Oreochromis niloticus

(Tilapia) fingerlings’ weights.

G. Analyzing the General Data using Statistical Tool

In order to analyze the general data, ANOVA (Analysis of Variance) will be

used to determine the potentiality of Lemna minor (Duckweed) meal and Glycine max

(Soybean) meal as protein alternative for fish meal in Oreochromis niloticus (Tilapia)

fingerlings formulated diet as well as to determine if there is a significant difference

among the treatments in terms of their protein content and growth effect in terms of

weight.

Group 1 Group2 Group 3 Group 4 Group 5 Group 6

x

 x 2

24
 Moreover, to evaluate if there is a significant relationship between the protein

concentration of the formulated fish meals and the growth of the Oreochromis

niloticus (Tilapia) fingerlings in terms of weight, Pearson Correlation Coefficient (r)

will be utilized.

x y (x-) (y-) (x-) (y-) (x-)2 (y-)2

Pearson’s r (correlation) Formula:

25
Conceptual Framework

Evaluation of Lemna minor (Common Duckweed) Meal and Glycine max

(Soybean) Meal as Potential Protein Alternative for Fish Meal in Oreochromis
niloticus (Tilapia) Fingerlings Formulated Diet

Independent Variable Dependent Variable

Lemna minor (Common Protein Alternative for

Duckweed) Meal and Glycine Fingerlings Formulated Diet
max (Soybean) Meal as
Formulated Diet

Figure 3 shows the independent and dependent variable. The independent

variable is the Lemna minor (Common Duckweed) Meal and Glycine max (Soybean)

Meal as Formulated Diet and the dependent variable is the Potential Protein

Alternative for Fingerlings Formulated Diet

26
 E. EXPECTED OUTPUT AND POTENTIAL IMPACT

1. Generally, the approximate crude protein content of Lemna minor (Duckweed)

plant on a dry weight is around 16%-45% (Zhao et al., 2014), whereas,

Glycine max (Soybean) has an average crude protein of 40-44% (Zhou et al.,

2018). With this, it can be assumed that the treatments will vary according to

these percentages and that they have the adequate protein concentration as fish

meal in Oreochromis niloticus (Tilapia) fingerlings formulated diet.

2. The results of the study of Asadi et al. (2012) reported that Lemna minor

(Duckweed) is more potent when it comes to its growth effect considering that

the protein concentration of Lemna minor (Duckweed) was generally rich in

leucine and lysine. Elaborately, the protein in Lemna minor (Duckweed) had

greater amino acid content than Glycine max (Soybean) protein. Amino acids

are commonly used for body upkeep, health, and the synthesis of new

structural proteins that will maximize feed efficiency and growth.

Furthermore, the study of Srirangam (2016) revealed that the most effective

when it comes to contributing the growth of the fish was the treatment with

20% Lemna minor (Duckweed) meal combined with 80% commercial fish

meal. With the data given, one of the possible alternative protein sources that

will be potentially more potent in terms of its growth (weight) effect is the

treatment wherein Lemna minor (Duckweed) meal and commercial fish meal

are combined together.

3. According to the findings of the study of Ovie and Eze (2013), the growth rate

of the fish was proportional to the degree of feed utilization efficiency; hence

high feed utilization efficiency resulted in relatively high growth rates. It was

27
 also found out that if the feed provides the proper number of essential amino

acids required by fish species, the optimal protein for the species will be met,

contributing to effective growth effect. Thus, it can be assumed that the

protein concentration and growth effect of each treatment are directly

proportional to each other.

4. According to the determined crude protein content of Lemna minor

(Duckweed) which is 16%-45% (Zhao et al., 2014) and Glycine max

(Soybean) which is 40-44% (Zhou et al., 2018) it can be assumed that there

will be no really significant difference in the evaluation of the potentiality of

alternative protein source among the treatments in terms of the protein

concentration.

5. Establishing the conclusion of Ovie and Eze (2013) as the basis that that the

protein concentration and growth effect of each treatment are directly

proportional to each other, it can be supposed that there will be no any

significant difference in the evaluation of the potentiality of alternative protein

source among the treatments in terms of growth effect considering the

assumption that there will be no significant difference among the treatments in

terms of the protein concentration of the alternative protein sources according

to Zhao et al. (2014) and Zhou et al. (2018).

28
 F. WORKPLAN AND TARGET DELIVERABLES

RESEARCH PROJECT TIMELINE

July 2022
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

Preparation Collection of Drying Procedures Pulveri- Proxi- Assem- Acclimati-

of Alternative zing mity bling Fish zing of fish Feeding Trial
Materials Protein Test Tanks
Sources

RESEARCH PROJECT TIMELINE

August 2022
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

Cleaning Statistical Analysis and Final

Feeding Trial and Documentation
Proper
Disposal

29
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32
 CURRICULUM VITAE

PERSONAL DATA:

Name: Dela Cruz, Allyza Kaye A.

Address: Prk.1 Brgy. Kinilis, Polomolok,

South Cotabato

Date of Birth: October 04, 2003

Place of Birth: Fatima, General Santos

City

Parents: Allan B. Dela Cruz

Emily A. Dela Cruz

Contact #: 09094247745

EDUCATIONAL BACKGROUND:

Elementary: Bonifacio R. Tagaban Sr. Integrated School

Prk.1 Brgy. Kinilis, Polomolok, South Cotabato

(2010-2016)

Junior High School: Polomolok National High School

Ocatvio Village, Cannery Site, Polomolok, South

Cotabato

(2016-2020)

Senior High School: Polomolok National High School

Ocatvio Village, Cannery Site, Polomolok, South

Cotabato

(2020-2022)

33
PERSONAL DATA:

Name: Fabulare, Leonah Jane S.

Address: Prk.1-A, Brgy. Landan

Polomolok, South Cotabato

Date of Birth: May 24, 2003

Place of Birth: Polomolok, South Cotabato

Parents: Jonathan D. Fabulare

Lorelyn S. Fabulare

Contact #: 09091732577

EDUCATIONAL BACKGROUND:

Elementary: Landan Elementary School

Brgy. Landan, Polomolok South Cotabato

(2010-2012)

Polomolok Central Elementary School

Polomolok, South Cotabatato

(2012-2014)

Dole Cannery Central Elementary School

Octavio Village, Cannery Site, Polomolok, South Cotabato

(2014-2016)

Junior High School: Poblacion Polomolok National High School

Poblacion, Polomolok, South Cotabato

(2016-2018)

Saint Lorenzo School of Polomolok

Prk. San Agustin, Polomolok, South Cotabato

34
 (2018-2020)

Senior High School: Polomolok National High School

Octavio Village, Cannery Site, Polomolok, South

Cotabato

(2020-2022)

35
PERSONAL DATA:

Name: Roderos, Jhamaica B.

Address: Prk. Faustino,

Javier, Poblacion,

Polomolok, South Cotabato

Date of Birth: July 16, 2003

Place of Birth: Polomolok, South

Cotabato

Parents: Lorine G. Roderos

Maria Nida C. Roderos

Contact #: 09515514019

EDUCATIONAL BACKGROUND:

Elementary: Upper Klinan Elementary School

Brgy. Upper Klinan, Polomolok, South Cotabato

(2010-2016)

Junior High School: Polomolok National High School

Octavio Village, Cannery Site, Polomolok, South

Cotabato

(2016-2020)

Senior High School: Polomolok National High School

Octavio Village, Cannery Site, Polomolok, South

Cotabato

(2020-2022)

36
PERSONAL DATA:

Name: Sandoy, Ellorvie C.

Address: Upper Maabtik, Esposado

Village Cannery Site,

Polomolok,

South Cotabato

Date of Birth: June 21, 2003

Place of Birth: Digos City, Davao

Parents: Elizalde E. Sandoy

Lorrefel C. Sandoy

Contact #: 09488891948

EDUCATIONAL BACKGROUND:

Elementary: Rizal Elementary School

Rizal St. Bansalan, Digos City, Davao Del Sur

(2010-2013)

Dole Cannery Central Elementary School

Octavio Village, Cannery Site, Polomolok, South Cotabato

(2013-2016)

Junior High School: Polomolok National High School

Octavio Village, Cannery Site, Polomolok, South

Cotabato

(2016-2020)

37
Senior High School: Polomolok National High School

Octavio Village, Cannery Site, Polomolok, South

Cotabato

(2020-2022)

38