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PRODUCTION OF SINGLE CELL PROTEIN FROM WATERMELON [CITRULLUS LANATUS (THUMB.) MATSUM AND NAKAI] WASTE USING ASPERGILLUS NIGER AND SACCHAROMYCES CEREVISIAE

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ABSTRACT

In this work, dry and wet peels derived from watermelon waste as alternative substrate for Single Cell Protein production from Saccharomyces cerevisiae (from palm wine), Aspergillus niger and Saccharomyces cerevisiae (from baker’s yeast) were investigated. Submerged method was employed to culture them in batches. A glucose solution (monosaccharide) was used as a control for the experiment. Batch 1 represent 4-day culture of the three microorganisms grown on dry, wet and control substrates respectively. Batch 1 gave rise to the following biomass in grams; for palm wine yeast – 0.10, 0.08 and 0.07; for baker’s yeast; 0.15, 0.35 and 0.32; for A. niger – 0.24, 0.30 and 0.21. Batch II represent 6-day culture of the microorganisms grown on dry, wet and control substrates respectively giving rise to the following biomass yield in grams; for palm wine yeast; 0.14, 0.35 and 0.15; for baker’s yeast – 0.33, 0.45 and 0.41; for A. niger – 0.26, 0.43 and 0.28. Batch III represent 8-day culture of the microorganisms grown on dry, wet and control substrates respectively which gave rise to the following biomass yield; for palm wine yeast – 0.10, 0.15 and 0.08; for baker’s yeast – 0.32, 0.43 and 0.29; A. niger – 0.13, 0.32 and 0.25. Nutritional compositions of the raw dry and wet watermelon waste prior to their use as substrates were determined. Proximate composition of the fungal biomass was also determined. Present investigation revealed that baker’s yeast grown on the wet watermelon substrate cultured for 6-day produced the highest quantity of single cell protein, suggesting that wet watermelon peel is more suitable for the production of SCP. The findings from this study support the need for SCP production from cheap, inexpensive agro-waste materials.





TABLE OF CONTENTS

Certification  .           .           .           .           .           .           .           .            .           .           i

Declaration    .           .           .           .           .           .           .           .            .           .           ii

Dedication      .           .           .           .           .           .           .           .            .           .           iii

Acknowledgement    .           .           .           .           .           .           .            .           .           iv

Table of contents       .           .           .           .           .           .           .            .           .           v

List of Tables and Figures   .           .           .           .           .           .            .           .           vii

Abstract         .           .           .           .           .           .           .           .            .           .           viii


CHAPTER ONE                                                            

1.0       Introduction .           .           .           .           .           .           .            .           .           1

1.1       Fungi .           .           .           .           .           .           .           .            .           .           2

1.2       Botany of Watermelon [Citrullus lanatus (Thumb.) Matsum

and Nakai]     .           .           .           .           .           .           .            .           .           3

1.3       Aims and Objectives             .           .           .           .           .            .           .           4

1.4       Justification   .           .           .           .           .           .           .            .           .           4       
   

CHAPTER TWO

2.0       Literature Review     .           .           .           .           .           .            .           .           6

2.1       Choice of Microorganism for SCP Production      .           .            .           .           8

2.2       Potential Substrates for Single Cell Protein Production .            .           .           11

2.3       Cultivation Methods in SCP Production    .           .           .            .           .           12

2.4       Nutritional Value of SCP     .           .           .           .           .            .           .           15

2.5       The Biotechnology of SCP Production       .           .           .            .           .           16

2.5.1    The Biochemistry and Physiology of Biomass Production             .           .           16

2.5.2    Process Design and Control             .           .           .           .            .           .           18

2.6       The Economics of SCP Production and Marketing          .            .           .           20

2.7       Drawbacks of Single Cell Protein Technology      .           .            .           .           20

2.8       Future Prospects       .           .           .           .           .           .            .           .           22


CHAPTER THREE

3.0       Materials and Methods        .           .           .           .           .            .           .           23

3.1       Collection of the Plant Material      .           .           .           .            .           .           23       

3.2       Preparation of the Plant Sample     .           .           .           .            .           .           23

3.2.1    Dry Sample    .           .           .           .           .           .           .            .           .           23

3.2.2    Wet Sample   .           .           .           .           .           .           .            .           .           23

3. 3      Preparation of Watermelon Extracts         .           .           .            .           .           23

3.4       Microorganisms Used           .           .           .           .           .            .           .           25

3.5       Analysis of the Raw Substrate         .           .           .           .            .           .           25

3.6       Production and Harvesting of Single Cell Protein (SCP) .            .           .           25

3.7       Analysis of Fungal Biomass and Procedures for

Proximate Composition        .           .           .           .           .            .           .           26

3.7.1    Moisture Content      .           .           .           .           .           .            .           .           26

3.7.2    Determination of Ash Content        .           .           .           .            .           .           26

3.7.3    Determination of Crude Fat Content (Dry Extraction)    .            .           .           27

3.7.4    Determination of Crude Fat Content (Wet Extraction)   .            .           .           28

3.7.5    Determination of Crude Protein     .           .           .           .            .           .           28

3.8       Procedure for Total Sugar using Anthrone Reagent        .            .           .           29


CHAPTER FOUR

4.0       Results            .           .           .           .           .           .           .            .           .           31


CHAPTER FIVE

5.0       Discussion and Conclusion .           .           .           .           .            .           .           40

5.1       Discussion      .           .           .           .           .           .           .            .           .           40

5.2       Conclusion     .           .           .           .           .           .           .            .           .           42

REFERENCES         .           .           .           .           .           .           .            .           .           43

 

 

 

 

 

 

LIST OF TABLES AND FIGURES

Figure 1:        Proximate composition of the raw dry and wet watermelon peels prior to Fermentation               .           .           33

 

Figure 2:        Biomass (g) growth of fungi grown on two different substrates (dry and wet watermelon and glucose sugar as control) harvested at 4, 6 and 8 days after inoculation.           .           .           .           34

 

Figure 3:        Ash content of fungal biomass         .       .            .           .           35

 

Figure 4:        Crude fat content of fungal biomass           .           .            .           .           36

 

Figure 5:        Moisture content of fungal biomass            .           .            .           .           37

 

Figure 6:        Crude protein content of fungal biomass   .           .            .           .           38

 

Table 1:          Comparism of Microorganisms for Mean Biomass

Yield and Proximate Composition  .           .           .            .           .           39

 

Table 2:          Correlation Coefficient (r) of Biomass and Proximate Compositions

of Microorganisms Inoculated on Different Media            .           .           39

 

Table 3:          Comparison of Means of Raw Wet and Dry Watermelon Peel

Substrate used for Proximate Analysis       .           .            .           .           39

 

 


 

 

CHAPTER ONE

INTRODUCTION

The rapidly increasing world population necessitates the challenge of providing necessary food sources. The increasing global protein deficiency in food consumed by many people is becoming a major problem for mankind (Gour et al., 2015). Single Cell Protein (SCP) is one of the most important steps for this goal and is an alternative and an innovative way to successfully solving the global protein-rich food problem (Najafpur and Ghasem, 2007). With the world population reaching 9 billion by 2050, there is strong evidence that agriculture alone will not be able to meet the world’s food demand (Challinor et al., 2014). Proteins are mainly called as building blocks of all living organisms. Proteins are necessary for the growth and development of living organisms and carryout a number of different biochemical reactions in the form of enzyme (Khan et al., 2010). As compared to other macromolecules, our body requires abundant quantity of proteins and its destruction is sometimes lethal (Solomons, 1983). The protein comes from a number of vegetables, cereals and fruits, often not affordable by a common man and therefore microbial proteins can be an alternative source to make proteinous food available to the down trodden communities in the world in general and Nigeria in particular (Mahnaaz et al., 2010).

The term Single Cell Protein (SCP) refers to dead, dry microbial cells or total proteins extracted from pure microbial cell culture and is produced using a number of different microorganisms including bacteria. Fungi and algae (Anupama and Ravindra, 2000). Single Cell Protein can also be referred to as edible unicellular microorganisms. The biomass or protein extract from pure or mixed cultures of algae, yeasts, fungi or bacteria, may be used as an ingredient or a substitute for protein-rich foods and is suitable for human consumption or as animal feeds (https://wikipedia/single-cell-protein).

The benefits of Single Cell Protein were thus extended from the production of food to the preservation of the environment. It is an established fact that despite manifold increase in organization and industrialization, majority of our people live below poverty line and are malnourished and deserve immediate attention of the relevant government authorities in general and scientists in particular for ensuring that these people are with balanced food and nutritive diet, which comprises of all necessary ingredients including proteins which comprises of all necessary ingredients including proteins which are desperately needed for their healthy growth and development (Khan et al., 2010). Moreover, there is problem of disposal of organic wastes mainly consisting of vegetable and fruit residues. It is therefore considered imperative to make use of these affordable cheap substrates for culturing fungal species which can be harvested as a source of Single Cell Protein to be used for human and animal consumption or as supplement in the food and fodder. This will enable at least to some extent to help malnourished and to minimize the load of pollution. Thus, if the waste is utilized regularly and systematically for the production of SCP, it will be a source of protein at affordable cost (Mahnaaz et al., 2010). Single Cell Protein appears to be the only feasible approach to bridge the gap between requirement and supply of proteins (Singh, 1998).

Single Cell Protein was introduced by Carl Wilson in 1944 for the first time. Single Cell Protein in terms called cells or proteins is derived from microorganisms “microbial protein”, resulting from the growth of microorganisms on different sources of protein and carbohydrates was produced and used as an alternative or supplementary source of food in

the diet of animals and humans (Jamel et al., 2007; Adedayo et al., 2011).


1.1       FUNGI

Saccharomyces cerevisiae is a species of yeast. It has been instrumental to winemaking, baking, and brewing since ancient times (Walker et al., 2004). Many proteins important in human biology were first discovered by studying their homologs in yeast; these proteins include cell cycle proteins, signaling proteins, and protein-processing enzymes. S. cerevisiae is currently the only yeast cell known to have Berkeley bodies present, which are involved in particular secretary pathways (Moyad, 2008). All strains of S. cerevisiae can grow aerobically on glucose, maltose, and trehalose and fail to grow on lactose and cellobiose. However, growth on other sugars is variable. Galactose and fructose are shown to be two of the best fermenting sugars. The ability of yeasts to use different sugars can differ depending on whether they are grown aerobically or anaerobically (Feldmann, 2010).

Baker’s yeast (Saccharomyces cerevisiae) was the first microorganism to be produced in aerobic stirred fermentation on molasses as it is still produced today (White, 1954; Chen and Chinger, 1985).

Aspergillus niger is a fungus and one of the most common species of the genus Aspergillus (Samson et al., 2001). A. niger is cultured for the industrial production of many substances. Various strains of A. niger are used in the industrial preparation of citric acid (E330) and gluconic acid (E574) and have been assessed as acceptable for daily intake by the World Health Organization. A. niger fermentation is "generally recognized as safe" (GRAS) by the United States Food and Drug Administration under the Federal Food, Drug, and Cosmetic

Act ("Inventory of GRAS Notices: Summary of all GRAS Notices", 2008).


1.2       BOTANY OF WATERMELON [Citrullus lanatus (Thumb.) Matsum and Nakai]

Many raw materials have been considered as substrate (carbon and energy sources) for single cell protein production (Nasseri et al., 2011).

 Scientific Classification:

 Domain          -           Eukarya          

Kingdom         -           Plantae

Division          -           Magnoliophyta

Class               -           Magnoliopsida

Order               -           Cucurbitales

Family             -           Cucurbitaceae

Genus             -           Citrullus

Species            -           C. lanatus Thumb.

Generic Group            -           Gourd                          Source: (https://wikipedia.org)

Watermelon (Citrullus lanatus var. Lanatus, family; Cucurbitaceae) is a vine like flowering plant that originated from Southern Africa. It is large annual plant with coarse, hairy pinnately-lobed leaves with white to yellow flowers. It is grown for its edible fruit. Watermelon is a special kind of berry botanically called as “pepo”. The fruit has a smooth hard rind, usually green with dark green stripes or yellow spots and juicy, sweet interior flesh, usually deep red to pink but sometime orange, yellow or white, with many seeds. The fruit can be eaten raw or cooked (https://wikipedia.org).

A variety of microorganisms and substrates are used to produce SCP. Yeast is suitable for single cell protein production because of its superior nutritional quality (Miller and Litsky,

1976).


1.3       AIMS AND OBJECTIVES

1.         To determine the growth of selected microorganism (fungi) on waste watermelon peel (wet and dry forms) as carbon source (substrate) for SCP production.

2.         To determine which of the state of the substrate’s (watermelon) forms that will promote the growth of the organism better for the production of SCP.

3.         To determine at what period the highest yield will be obtained as well as the

microorganisms that will give the highest biomass yield.


1.4       JUSTIFICATION

Increasing human’s search for nutrition has led to increase in consumption of fruits and vegetables thereby leaving the environment polluted with agro-wastes from these fruits and vegetables. These wastes are found dumped on road sides, in the drains and water bodies proper. Disposal of wastes especially those from these fruits and vegetables (agro-wastes) has been a serious problem and their deposition poses health hazards for humans. Hence, there is urgent need for proper exploitation of these wastes as a substrate for the production of cellular biomass of edible protein (single cell protein) to supplement the scarce protein needed to serve as animal feed and food for human consumption. This single cell protein serves as suitable alternative to scanty protein sources through recycling of these wastes. The process of production is also cheap or inexpensive and environmentally friendly.

The present economic crisis in Nigeria today is posing a tremendous threat on livelihood of many citizens. The burgeoning population is faced with the challenge of proper feeding as majority become victim of malnutrition. Protein being one of the essential components of human and animal diet, the sources is very expensive for an average citizen of Nigeria and other under developed countries of the world. The innovation of single cell protein production as a means of obtaining a protein source from waste agricultural produce is a welcome development to help address the problem of protein need to humans and animals. Therefore, single cell protein becomes the only feasible approach to bridge the gap between requirement and supply of protein.

It is to this end that this research study is aimed at investigating the possibility of using dry and wet forms of watermelon peels as substrate/carbon source for production of single cell protein using three selected fungal organisms.

 

 

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