Amylolytic Activity Of Lactic Acid Bacteria Isolated From Nunu (A Dairy Product)

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