screening, culture optimization and utilization of marine diatom amphora coffeaeformis for ...

214 Pages·2015·7.46 MB·English

SCREENING, CULTURE OPTIMIZATION AND UTILIZATION OF
MARINE DIATOM AMPHORA COFFEAEFORMIS FOR SUSTAINED
PRODUCTION OF BIOFUEL AND AS AN AQUAFEED ADDITIVE

A thesis submitted to Bharathidasan University, Tiruchirappalli
in partial fulfillment of the requirements for the award of the Degree of
DOCTOR OF PHILOSOPHY
IN
MARINE SCIENCE

By
Ms. S. JEYANTHI

Under the supervision of
Dr. P. SANTHANAM

DEPARTMENT OF MARINE SCIENCE
SCHOOL OF MARINE SCIENCES
BHARATHIDASAN UNIVERSITY
TIRUCHIRAPPALLI-620 024.
TAMIL NADU, INDIA

DECEMBER 2015
CERTIFICATE
This is to certify that the thesis entitled “SCREENING, CULTURE
OPTIMIZATION AND UTILIZATION OF MARINE DIATOM AMPHORA
COFFEAEFORMIS FOR SUSTAINED PRODUCTION OF BIOFUEL AND AS
AN AQUAFEED ADDITIVE” submitted by Ms. S. JEYANTHI for the award of the
Degree of DOCTOR OF PHILOSOPHY IN MARINE SCIENCE is based on the result
of studies carried out by her under my supervision and guidance in the Department of
Marine Science, School of Marine Sciences, Bharathidasan University, Tiruchirappalli-24,
Tamil Nadu, India. This thesis or any part of this thesis has not been submitted elsewhere
for the award of any other degree/diploma of this or any other university.

Signature of the Guide

Date:
Place:
DEPARTMENT OF MARINE SCIENCE
Bharathidasan University
Tiruchirappalli, Tamil Nadu, India-620 024.

S. JEYANTHI [email protected]
Research Scholar (FT)

DECLARATION
The research work presented in this dissertation entitled “SCREENING,
CULTURE OPTIMIZATION AND UTILIZATION OF MARINE DIATOM
AMPHORA COFFEAEFORMIS FOR SUSTAINED PRODUCTION OF BIOFUEL
AND AS AN AQUAFEED ADDITIVE” has been carried out by me under the guidance
of Dr. P. SANTHANAM, Assistant Professor, Department of Marine Science, School of
Marine Sciences, Bharathidasan University, Tiruchirappalli-24. This work is original and
has not been submitted in part or full for any degree/diploma of this or any other
university.

Date: (S. JEYANTHI)
Place:
ACKNOWLEDGEMENTS
First and foremost I am indebted to my parents and sister who supported me in
making this long-term research endeavor a success.
I express my heartfelt gratitude to my research supervisor Dr. P. Santhanam,
Assistant Professor, Department of Marine Science, Bharathidasan University,
Tiruchirappalli, for providing me the opportunity to pursue my academic skills under his
guidance and expertise. Without his able guidance my Ph.D work would not have
happened, literally. His passion for marine biology is both inspirational and infectious.
I extend my appreciation to my doctoral committee members Dr. N. Thajuddin,
Professor and Head, Department of Microbiology, Bharathidasan University, Tiruchirappalli
for providing me the confocal microscopy facility and Dr. J. Sesh Serebiah,
Assistant Professor, Department of Marine Studies and Coastal Resource Management,
Madras Christian College, Tambaram, Chennai-54 for their suggestions and valuable
inputs in the development of my Ph.D work.
I would like to thank Emeritus Professors of Department of Marine Science
Dr. A. Rajendran and Dr. V. Radhakrishnan, BARD, Tiruchirappalli. I am forever
indebted to the kindness and hospitality of all the faculty members, Dr. R. Arthur James,
Asst. Professor & Head, Dr. R. Rajaram, Dr. N. Manoharan, and Dr. V. Yogananthan,
Department of Marine Science, Bharathidasan University, Thiruchirappalli for their
support since the beginning of my research in the department.
On this note, I would like to dedicate a special thanks to Dr. A. Antony Joseph
Velanganni, Assistant Professor, Department of Biochemistry and Dr. R. Renganathan,
Professor, Department of Chemistry, Bharathidasan University, Thiruchirappalli for
helping me with the fluorescence studies.
I owe a great deal to Dalmia Research Centre, Coimbatore for carrying out
HPTLC amino acid analysis and Mr. S. Kumaravel, Quality manager, IICPT, Thanjavur
for his contribution in GC-MS study.
My deepest thanks go to Dr. Mrs. Florida Tilton, Biozone Research Technologies,
Chennai for her assistance in Molecular taxonomy work.
I am grateful to Dr. A. Balamurugan, Assistant Professor, and Ms. S. Viji,
Research Scholar, Dept of Nanoscience and Technology, Bharathiar University,
Coimbatore for their contribution in the SEM studies.
In particular, a special thanks to my friends Dr. K. Maniselvan, Dr. A. Shenbaga
Devi, Dr. S. Dinesh Kumar, Mr. C. Prem Kumar, Dr. S. Ananth, Dr. B. Balaji
Prasath and Dr. R. Nanda Kumar for helping and supporting me with laboratory work
and preliminary studies.
I would like to thank my junior friends Ms. P. Nithya, Ms. P. Bharatha Rathna,
Ms. T. Jayalakshmi, Mr. M. Kaviyarasan, Mrs. R. Sasirekha, Ms. M. Sathiya Deepika,
Ms. P. Priyadarshini, Mr. M. Santhosh Gokul, Mr. T. Dinesh Kumar, Mr. R. Viji
and Mr. U. Devan for their assistance in several aspects of the work at all times.
Special thanks to Mr. S. Vinoth Kumar, Mr. V. Ramalingam, Mr. G. Rajiv
Gandhi, Ms. P. Chitra Devi and Mr. P. Vijayakumar for having so kindly and
promptly lent their laboratory space and equipment.
I would like to thank the Post Doctoral Fellow’s Dr. T. Veeramani,
Dr. S. Balakrishnan and Dr. N. Manickam for their cooperation during my study period.
I thank Mr. Jayaraj, Senior Technician, Rank Marine Hatchery, Pondicherry, for
providing shrimp seed for my experiment. My sincere thanks to Mr. C. J. Eswaran,
Technical manager, CLASWIN Aqua consultants Pvt. Ltd, Chennai for providing me
with commercial shrimp feeds.
Special thanks goes to my uncle Dr. George Abraham, Assistant Principal,
American College, Madurai for his inspirations and encouragement.
I am grateful to my friends Mr. A. Muruga Das and Mr. P. Balaji from MGDC,
Bharathidasan University, Tiruchirappalli for their assistance with fluorescence
microscopy work. I also thank my school friend Mr. A.C Ramesh Babu, Senior
Research Fellow, Department of Chemistry, Anna University, Tiruchirappalli for helping
me in biofuel property analysis.
Finally, I express my most humble gratitude to my best friend Mrs. V. Shyamala,
Research Scholar, C.A.S in Botany, University of Madras, who, despite the distance,
never failed to keep in touch with and had been a great moral support throughout my life.

S. JEYANTHI
I dedicate this thesis to my family and friends, who
have been, is & always will be my true source of
support & strength.
CONTENTS
CHAPTER PAGE
TITLE
NO. NO.
CHAPTER I 1. GENERAL INTRODUCTION 1
2. REVIEW OF LITERATURE 5
3. SCOPE AND OBJECTIVES 21
4. DESCRIPTION OF THE STUDY AREA 23
CHAPTER II 5. COLLECTION AND PRELIMINARY SCREENING OF 25-35
HIGH LIPID YIELDING MARINE DIATOMS
5.1 Introduction 25
5.2 Materials and Methods 27
5.3 Results 29
5.4 Discussion 33
CHAPTER III 6. COLLECTION, ISOLATION AND MOLECULAR 36-52
IDENTIFICATION OF MARINE DIATOMS
6.1 Introduction 36
6.2 Materials and Methods 38
6.3 Results 44
6.4 Discussion 51
CHAPTER 1V 7. OPTIMIZATION OF CULTURE CONDITIONS TO 53-84
ACHIEVE MAXIMIZED BIOMASS AND HIGH
LIPID YIELD
7.1 Introduction 53
7.2 Materials and Methods 55
7.3 Results 58
7.4 Discussion 82
CHAPTER PAGE
TITLE
NO. NO.
CHAPTER V 8. OPTIMIZATION OF BIOMASS CONDITION, 85-98
EXTRACTION METHODS AND SOLVENTS
CONDITIONS ON BIOFUEL YIELD
8.1 Introduction 85
8.2 Materials and Methods 87
8.3 Results 91
8.4 Discussion 96
CHAPTER VI 9. MASS CULTURE OF DIATOM AMPHORA 99-110
COFFEAEFORMIS USING AQUACULTURE
WASTEWATER
9.1 Introduction 99
9.2 Materials and Methods 101
9.3 Results 103
9.4 Discussion 108
CHAPTER VII 10. EXPLORATION OF CO-PRODUCTS FOR ZERO 111-135
WASTE MANAGEMENT
10.1 Introduction 111
10.2 Materials and Methods 113
10.3 Results 124
10.4 Discussion 133
11. SUMMARY AND CONCLUSION 136-138
REFERENCES 139-176
LIST OF ABBREVIATIONS
% - Percentage
ppm - Parts per million
PSU - Practical salinity unit
µl - micron litre
µm - micron meter
µg/ml - micron gram per milliliter
g/L - gram per liter
°C - Degree Centigrade
O.D - Optical Density
AU - Arbitrary Unit
LD - Lipid Droplet
LEA - Lipid extracted algae
GCMS - Gas Chromatograph Mass Spectrometer
nm - nano meter
% dcw - Percentage dry cell weight
FAME - Fatty Acid Methyl Esters
ANOVA - Analysis of Variance
DIC - Differential interference contrast
HPTLC - High performance thin layer chromatography
SEM - Scanning Electron Microscope
BLAST - Basic local alignment search tool
NCBI - National centre for biotechnology information