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ISOLATION AND SCREENING OF STREPTOMYCES SPECIES FOR ENHANCED AMYLASE PRODUCTION BY Martina NATHANIEL DEPARTMENT OF MICROBIOLOGY, FACULTY OF LIFE SCIENCES, AHMADU BELLO UNIVERSITY, ZARIA, NIGERIA JULY, 2017 i ISOLATION AND SCREENING OF STREPTOMYCES SPECIES FOR ENHANCED AMYLASE PRODUCTION BY MARTINA NATHANIEL [B.Sc. (Hons) Microbiology, A.B.U – 2011] (M.Sc./SCI/22064/2012-2013) ADISSERTATION SUBMITTED TO THE SCHOOL OF POSTGRADUATE STUDIES, AHMADU BELLO UNIVERSITY, ZARIA IN PARTIAL FULFILLMENT OF THE REQUREMENTS FOR THE AWARD OF A MASTER DEGREE IN MICROBIOLOGY DEPARTMENT OF MICROBIOLOGY, FACULTY OF LIFE SCIENCES, AHMADU BELLO UNIVERSITY, ZARIA NIGERIA JUNE, 2017 ii DECLARATION I declare that the work in this dissertation entitled “Isolation and screening of Streptomycesspp. for enhanced amylase production”has been carried out by me in the Department of Microbiology, Ahmadu Bello University, Zaria. The information derived from the literature has been duly acknowledged in the text and a list of references provided. No part of this dissertation was previously presented for another degree or diploma at this or any other institution. ____________________ _______________ Martina NATHANIEL Date iii CERTIFICATION This dissertation entitled “ISOLATION AND SCREENING OF STREPTOMYCESSPPFOR ENHANCED AMYLASE PRODUCTION”by Martina NATHANIEL meets the regulations governing the award of the degree of Master of Science in Microbiology of the Ahmadu Bello University, and is approved for its contribution to knowledge and literary presentation. Prof. S.A. Ado ________________________ _________________ ________________ Chairman, Supervisory Committee Signature Date Prof. I.O. Abdullahi ________________________ __________________ ________________ Member, Supervisory Committee Signature Date Prof. I.O. Abdullahi ________________________ ____________________ ________________ Head of Department Signature Date Prof. S.Z. Abubakar _________________________ ____________________ ________________ Dean, School of Postgraduate Studies Signature Date iv ACKNOWLEDGEMENT I give thanks to Almighty God for His faithfulness and for granting me the grace to carry out this research work. My gratitude also goes to my distinguished team of supervisors: Professors S.A. Ado and I.O. Abdullahi for their great support, guidance and making sure that the work is a success. I am so grateful and may God bless you beyond your expectation. I also want to acknowledge the following people, all of Department of Microbiology for their wonderful contributions and advice: Dr. A.B Suleiman, Dr. E.E. Ellah, Mallam Shuaibu and my wonderful course mates: Mrs. Safiya Aliyu and Mr. Ashiru Rasheed, may God reward you all. I also want to thank my lovely parents: Mr. Nathaniel Jibrin and Mrs. Helen Nathaniel for all the sacrifice they made in making sure that this work is successful. You will live to enjoy the fruit of your labor by God‟s grace. My appreciation also goes to my siblings: Angela Nathaniel, Edward Nathaniel, Victor Nathaniel, Sonia Nathaniel, Winifred Nathaniel and Wilfred Nathaniel for their encouragement, I am blessed to have you all. v DEDICATION This piece of work is dedicated to God Almighty for His faithfulness and divine favour granted me, which has enabled me to finish this work successfully. To God alone is all the Glory. vi ABSTRACT In this work, optimization of culture conditions for enhanced amylase production by five Streptomyces strains isolated from the agricultural soil from Ahmadu Bello University dam were investigated. Streptomyces isolates were identified by Gram staining, morphological and physiological observations and those identify were; Streptomyces narbonensis, Streptomyces michiganensis, Streptomyces orientalis, Streptomycesspp (S4), Streptomyces spp (S5). Upon screening for amylase production, only Streptomyces michiganensis was able to produce amylase with diameter of clear zones of starch hydrolysis of 38.3±0.45mm, hence it was used for further production of amylase under solid state fermentation. Determination of parameters such as period of fermentation, pH, temperature, moisture content and inoculum size were also carried out in this study. Highest amylase produced was observed at 72h incubation period with amylase activity of 8.5±21U/ml and 12.1±0.35U/ml using wheat bran and soluble starch respectively. The optimum period of fermentation, pH, temperature, initial moisture content and inoculum size were 48h (17.7±0.41U/ml), pH 6 (12.2±0.28U/ml), 35°C (11.2±1.13U/ml), 30% (7.9±0.21U/ml) and 2ml of 1.0 x106spores/ml (7.9±0.21U/ml) respectively. There was a significant increase in enzyme production after optimization of fermentation condition, with maximum yield of 17.7±0.41U/ml recorded at 48h.Wheat bran could serve as a good substrate for amylase production by Streptomyces michiganensis. vii TABLE OF CONTENTS Title page--------------------------------------------------------------------------------------------------------i declaration ...................................................................................................................................... iii Certification ................................................................................................................................... iv Acknowledgement .......................................................................................................................... v Dedication ...................................................................................................................................... vi Abstract ......................................................................................................................................... vii Table of Contents ......................................................................................................................... viii List of Tables ................................................................................................................................ xii List of Figures .............................................................................................................................. xiii List of Plates ................................................................................................................................ xiv List of Appendices ........................................................................................................................ xv CHAPTER ONE ........................................................................................................................... 1 1.0 Introduction ............................................................................................................................ 1 1.1 Background of the Study .................................................................................................... 1 1.2 Statement of Research Problem ......................................................................................... 3 1.3 Justification of the Study .................................................................................................... 4 1.4 Aim of the Study .................................................................................................................. 4 1.5 Objectives of the Study ....................................................................................................... 5 CHAPTER TWO .......................................................................................................................... 6 2.0 Literature Review .................................................................................................................. 6 2.1 Actinomycetes ...................................................................................................................... 6 2.2 Genus Streptomyces ............................................................................................................ 6 viii 2.3The Life Cycle of Streptomycetes ....................................................................................... 7 2.4 Streptomycetes Habitats ......................................................................................................8 2.4.1 Hay and other Organic Material ..................................................................................... 8 2.4.2 Fresh Water and Marine Habitats ................................................................................... 9 2.4.3 Plants .............................................................................................................................. 9 2.4.4 Animal and Humans ..................................................................................................... 10 2.5 Growth Requirements of Streptomycetes ....................................................................... 10 2.6 Metabolites Produced by Streptomycetes ....................................................................... 10 2.7 Usefulness of Streptomyces spp. In Biotechnology .......................................................... 11 2.8 Amylase .............................................................................................................................. 12 2.9 Classification of Amylase .................................................................................................. 12 2.9.1 α-Amylase ..................................................................................................................... 12 2.9.2 β-Amylase ....................................................................................................................
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