ABSTRACT

Amylolytic activity of Lactic acid bacteria isolated from Nunu (a dairy product) was studied. Lactic acid bacteria were isolated from Nunu obtained from three sources and identified accordingly. Amylolytic activity of the LAB isolate were evaluated under varying conditions of temperature, pH and incubation time for optimization purposes. Results obtained show that three species of Lactic acid bacteria were isolated from the Nunu with varying levels of occurrences including Lactobacillus fermentum (80%), Lactobacillus pentosus (60%), and Pediococcus acidilatici (60%). All these isolates demonstrated amylolytic activities creating halo zones on starch containing medium with diameters of 5.03mm, 4.33mm and 3.8mm for Lactobacillus fermentum, Lactobacillus pentosus and Pediococcus acidilatici respectively. Also the optimum growth conditions for enzyme (amylase) activity of the three LAB isolates shows pH 7.0 with amylase activity of 5.0mm halo zone size, pH 6.0 with amylase activity of 4.57mm halo zone size and pH 7.0 with amylase activity of 4.0mm halo zone size for the same isolates respectively. Optimum incubation temperatures of 50°C with amylase activity of 4.75, 60°C with amylase activity of 4.57 and 60°C with amylase activity of 4.43 also for the same isolates respectively. The optimum incubation time was 72hrs for Lactobacillus fermentum, 72hrs for Lactobacillus pentosus and 96hrs for Pediococcus acidilatici at which the respective amylase activities were 4.9mm, 4.57mm and 4.33mm halo zone sizes respectively. It was observed that there were variations in the type and occurrence of LAB in Nunu while their optimal growth activities varied slightly but significantly.

TABLE OF CONTENTS

Title page i

Certification ii

Dedication iii

Acknowledgements iv

Table of Contents v

List of Charts viii

List of Tables ix

Abstract x

CHAPTER ONE

1.0   Introduction 1

1.1   Aims And Objectives 2

CHAPTER TWO

2.0   Literature Review 4

2.1   History 4

2.2   Human Evolution 5

2.3   Alpha-amylase 6

2.4   Beta-amylase 7

2.5   Gamma-amylase 7

2.6   Uses 7

2.6.1   Fermentation 7

2.6.2   Flour Additive 8

2.6.3   Molecular Biology 9

2.6.4   Medical Uses 9

2.6.5   Other Uses 9

2.7   Hyperamylasemia 10

2.8   Microorganisms and Amylase 10

2.9   Production of Amylase 12

CHAPTER THREE

3.0   Materials and Methods 14

3.1   Materials/Equipment 14

3.2   Sources of Materials (Sample Collection) 14

3.3   Sample/Media Preparation 14

3.4   Isolation of Lactic Acid Bacteria from Nunu 15

3.4.1   Isolation of Lactic Acid Bacteria 16

3.4.2   Characterization of Lactic Acid Bacteria Isolates 17

3.4.2.1   Colony Characteristics 17

3.4.2.2   Microscopic Characteristics 18

3.4.3   Biochemical Tests 18

3.4.3.1   Catalase Test 18

3.4.3.2   Citrate Test 19

3.4.3.3   Motility Test 19

3.4.3.4   Oxidase Test 19

3.4.3.5.  Voges Proskauer (V.P) Test 20

3.4.4.1   Growth at Different Temperatures 20

3.4.4.2   Salt Tolerance Test 20

3.4.4.3   Carbohydrate Fermentation Test 20

3.5   Determination of Amylase Activity 21

3.6.0   Optimization of Growth Factors for Amylase Activity 21

3.6.1   Optimization of Temperature for Amylase Production 21

3.6.2   Optimization of pH for Amylase Production 22

3.6.3   Optimization of Incubation Time 22

CHAPTER FOUR

4.0   Results 24

CHAPTER FIVE

5.0   Discussion 30

5.1   Conclusion 31

5.2   Recommendation 31

References 32