ISOLATION AND CHARACTERIZATION OF MICROORGANISMS RESPONSIBLE FOR THE FERMENTATION OF TIGERNUT MILK

ABSTRACT

The microorganisms responsible for the fermentation of tiger nut milk sold in Umuahia, Abia State were investigated. The isolation, identifications, characterization and microbial counts were done using standard microbiological techniques. The wet and dry tiger nut sample at zero fermentation period has microbial load range from 3.0 + 0.3d to 6.05 + 0.8d and 3.75 + 0.5e to 6.95 + 0.8c while at 72 hours of fermentation has 3.45 + 0.4d to 9.0 + 0.9b and 2.8 + 0.4d to 7.0 + 0.4b and at 120 hours of fermentation has 3.25 + 0.3c to 8.35 + 0.9d and 3.9 + 0.6c to 6.75 + 0.6e respectively. It reveals that microbial load increased from zero hour to 120 hours of fermented periods. The mean viable microbial counts in the samples analyzed were above the recommended standard. The predominant bacterial and fungal species includes; Bacillus subtilis, Escherichia coli, Klebseilla Species, Staphylococci aureus, Lactobacillus species, Lactobacillu Plantarum, Aspergillus flavus, Mucor species and Saccharomyces cerevisiae with varying degrees of occurrences. Escherichia coli has the highest number of isolates (24%) followed by Lactobacillus species (18.2%) while Aspergillus flavus has the least isolate with (4%). The results obtained in this study shows that there are presence of pathogenic bacteria that may be potential source of food borne infection and some related diseases for the consumers of this product in the studied areas. Therefore, further studies are recommended to preserve the health of the consumers.

TABLE OF CONTENTS

Declaration                                                                                                                              iii

Certification                                                                                                                            iv

Dedication                                                                                                                               v

Acknowledgements                                                                                                                vi

Table of contents                                                                                                                    vii

List of tables                                                                                                                           x

Abstract                                                                                                                                    xi 

CHAPTER ONE

1.0       Introduction                                                                                                                1

1.1       Aims and objectives                                                                                                   3

CHAPTER TWO

2.0     Literature Review                                                                                                       4

2.1       Nutritional Characteristics and Bioactive Compounds of Tiger Nut                                     6

2.2       Heath Benefits of Tiger Nut                                                                                      8

2.3       Fermentation and fermented products                                                                        9

2.4       Fermentation of Tiger Nut                                                                                          11

CHAPTER THREE

3.0       Materials and Methods                                                                                               14

3.1       Sample Collection                                                                                                      14

3.2       Preparation and Fermentation of Tiger nut Milk                                                        14

3.3       Microbiological Analysis                                                                                           14

3.4       Characterization of Isolates                                                                                        15

3.5       Identification of bacteria                                                                                            15

3.6       Catalase test:                                                                                                               16

3.7       Citrate test:                                                                                                                 16

3.8       Indole test:                                                                                                                  16

3.9       Methyl red test:                                                                                                           17

3.10     Urease test                                                                                                                  17

3.11     Identification of isolated fungi                                                                                   18

    CHAPTER FOUR

4.0       Results                                                                                                                        19                                            

CHAPTER FIVE

5.0       Discussion, Conclusion and Recommendation                                                          30

5.1       Discussion                                                                                                                   30

5.2       Conclusion                                                                                                                  33

5.3       Recommendation                                                                                                       34

References                                                                                                                  35

LIST OF TABLES

TABLES                    TITLE                                                               Page

4.1                   Mean viable microbial count of fermented dry tiger nut.                            20

4.2                   Mean viable microbial count of fermented dry tiger nut.                            22

4.3                   Identification and characterization of bacteria Isolated from two different

fermented tiger nut samples                                                                           24

4.4                   Identification and characterization of yeast and fungi from the two

different fermented tiger nut samples                                                             26

4.5.                  Distribution of microbial isolates from the two different fermented tiger

nut samples                                                                                                     28

4.6.                  Percentages occurrence of microbial isolates from the two different

fermented tiger nut samples.                                                                          30

LIST OF FIQURES

Fig 1: Flowchart showing Tigernut Milk                                                  13

CHAPTER ONE

1.0    INTRODUCTION

Milk has been recognised as an important food for infants and growing children (Obizoba, and  Anyika, 1995). In developing countries, the cost of dairy milk and their products is prohibitive and this has led to the development of alternative source of milk from plant materials (Singh, and Baius, 1988). An inexpensive milk substitute extracted from locally available plant foods like legumes with satisfactory quality and rich in protein could play an important role in protein malnutrition (Ukwuru, and  Ogbodo, 2011), source of producing acceptable nutritious drink (Hankus, and Sarret, 1967) and alleviate problem of short food supply (Reddy et al, 1990).

Tigernut (Cyperus esculenta), also known in Nigeria as “Ayaya” in Hausa, “Ofio” in Yoruba and “Akiausa” in Igbo has three varieties (black, yellow and brown) which are underutilized due to lack of information on their nutritional potentials (Omode et al, 1995).   It can be eaten raw, roasted, dried, baked or be made into milk (Oladele, and  Aina, 2007). Fermentation of tigernuts could produce varieties of milk products such as natural tigernut milk, pasteurized tigernut milk, sterilized tigernut milk, ultra-high temperature tigernut milk, concentrated and condensed tigernut milk (Ukwuru, and Ogbodo, 2011). Tigernut milk is a very nutritive and energetic drink both for old and young people (Abaejoh, et al, 2006), rich in energy content (starch, fat, sugars and protein), mineral (phosphorus, potassium) and vitamins E and C (Belewu and Belewu, 2007). It has enormous health benefits which include prevention of heart attacks, thrombosis, activation of blood circulation and reduction in the risk of colon cancer. It is also suitable for diabetic patients and helps in reducing weight. The nuts are soaked in water, wet milled, sieved, sweetened and flavoured. Soaking is a major unit operation because it can be used to reduce soluble antinutrients such as tannins and polyphenols. Sponteneous fermentation of tigernut milk results from the competitive activities of a variety of contaminating microorganisms. Those best adapted to the substrate and to technical control parameters, eventually dominate the process. The production of metabolites inhibitory to other contaminating organisms may provide additional advantage during fermentation (Holzapfel, 2000). Milk contains different species of microorganisms in varying numbers and their activity is uncontrolled, thus, uniformity of product is difficult to achieve during fermentation. It is often necessary and desirable to carry out controlled fermentation with specific organisms and this has led to the propagation and selection of known cultures, usually referred to as starter cultures. Similar to any other milk, principal organisms employed as starter include species of lactic acid bacteria including Streptococcus, Leuconostoc and Lactobacillus which may be a pure or mixed cultures. The proper selection and balance for starter culture are critical for the manufacture of fermented products of desirable texture and flavour. Quality and reproducibility of fermented milks and processes are ensured by using industrial starters, nevertheless, consumers prefer traditional fermented milks since artisanal starters give these products more typical flavours (Wouters, et al, 2002).

Tiger nut (Cyperus esculentus) is a tuber, mainly harvested in Spain, West Africa countries like Nigeria, Senegal, or Ghana, and also in South America, as in Chile. From an economic point of view, tiger nut is described as an underutilized African crop with high potential for development. Because of its ecological plasticity and its invasive capacity, this plant is considered as a weed or a crop depending on the context. In West Africa, the tubers are often part of the diet as they are cheap, available all the year around, and with nutritional benefits. (Yeboah et al, 2012)

1.1       AIMS AND OBJECTIVES

The aim of this research is to isolate and characterize the microorganisms responsible for the fermentation of tiger nut milk

The objectives are;

•       To isolate microorganisms (bacteria and fungi) responsible for the fermentation of tiger nut milk

•       To identify and characterise the isolated micro organisms

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