KWAME NKRUMAH UNIVERSITY OF SCIENCE AND TECHNOLOGY COLLEGE OF SCIENCE PROTEIN EXTRACTION FROM FERN AND ITS PHYSICOCHEMICAL PROPERTIES A THESIS SUBMITTED TO THE DEPARTMENT OF BIOCHEMISTRY AND BIOTECHNOLOGY IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE AWARD OF THE DEGREE OF MASTER OF PHILOSOPHY IN FOOD SCIENCE AND TECHNOLOGY BY EDWARD KEN ESSUMAN (BSC. AGRICULTURAL SCIENCE) JULY, 2013 CERTIFICATION PAGE I hereby declare that this submission is my own work toward the MPhil and that, to the best of my knowledge, it contains no material previously published by another person nor material which has been accepted for the award of any other degree of the University, except where due acknowledgement has been made in the text. EDWARD KEN ESSUMAN …………… ……………… (PG 5016610) Signature Date Certified by: MRS. GLORIA M. ANKAR-BREWOO ...…..……… …………… JOHN BARIMAH …………… ……………. (Supervisors) Signature Date Certified by: PROF. (MRS) I. ODURO …………… ……………… (Head of Department) Signature Date ii ABSTRACT Fern is a seedless vascular plant that reproduces via spores and has various usefulness. This study was carried out to optimize conditions for protein extraction from two defatted fern types namely Nephrolepis biserrata and Arthropteris orientalis. The extraction of defatted fern protein was conducted using alkaline, alcohol and saline treatments method under various conditions such as variation of treatments concentration and time of agitation. Central composite design of response surface methodology was used for identification of the best condition and extraction yield optimization. Result showed that alkaline treatment of 0.1 M NaOH and agitation time of 30 min produced the highest extraction yield with maximum protein recovery of 5.11 mg/mL and 2.03 mg/mL for Nephrolepis biserrata and Arthropteris orientalis respectively. Nephrolepis biserrata fern type gave relatively higher maximum protein yield, giving about more than twice the values obtained for the Arthropteris orientalis. Nephrolepis biserrata showed the highest foaming capacity (65%) and solubility (55.9%). For each isolate, water and oil holding capacity were not significantly different (p>0.05) with Nephrolepis biserrata recording 2.13 mL/g and 2.73 mL/g and Arthropteris orientalis recording 2.39 mL/g and 2.93 mL/g respectively. Protein types such as β-galactosidase, phosphorylase and lysozyme were found to be present in the fern protein isolates in terms of their molecular weight. iii TABLE OF CONTENTS TITLE PAGE…………………………………………………………………………i CERTIFICATION PAGE ....................................................................................................... ii ABSTRACT ........................................................................................................................... iii LIST OF TABLES ................................................................................................................ vii LIST OF FIGURES .............................................................................................................. viii ACKNOWLEDGEMENT ..................................................................................................... ix DEDICATION ........................................................................................................................ x CHAPTER ONE ..................................................................................................................... 1 1.0 Introduction ....................................................................................................................... 1 1.1 Statement of the Problem .............................................................................................. 4 1.2 Justification of Work ..................................................................................................... 4 1.3 Goal ............................................................................................................................... 6 1.3.1 Objective ................................................................................................................ 6 CHAPTER TWO ..................................................................................................................... 7 2.0 LITERATURE REVIEW .................................................................................................. 7 2.1 Introduction ................................................................................................................... 7 2.2 Evolution and Classification of Fern ............................................................................. 8 2.3 Distribution and Abundance of Fern ............................................................................. 8 2.4 Uses of Indigenous Edible Wild Plant ........................................................................ 10 2.4.1 Effect of processing temperature on nutrients of indigenous leafy vegetable ...... 11 2.5 Fern Consumption and Utilization .............................................................................. 11 2.6 Preservation of Fern .................................................................................................... 15 2.7 Nutritional Components of Fern.................................................................................. 16 2.7.1 Nutritional analysis of fern ................................................................................... 17 2.7.2 Nutritional determination of sporangia and leaf of fern ....................................... 17 2.7.3 Total amino acid compositions of tropical ferns .................................................. 19 2.7.4 Factors that inhibit nitrogen content in some fern species ................................... 24 2.7.5 Physicochemical and functional properties of fern fronds and rhizome .............. 24 2.8 Role of Protein ............................................................................................................ 25 2.9 Protein Assessment ..................................................................................................... 26 2.10 Protein Sources .......................................................................................................... 26 2.10.1 Animal source..................................................................................................... 27 2.10.2 Plant source ........................................................................................................ 27 2.11 Effects of Animal and Vegetable Protein .................................................................. 28 iv 2.12 Properties of Proteins in Food Systems ..................................................................... 29 2.12.1 Factors affecting properties of proteins in food systems .................................... 30 2.13 Chemical and Physical Properties of Food Proteins .................................................. 32 2.13.1 Molecular forces affecting physicochemical and functional properties ............. 33 2.14 Response Surface Methodology ................................................................................ 33 2.15 Extraction and Isolation of Proteins .......................................................................... 35 2.15.1 Factors that affect protein extraction .................................................................. 35 2.16 Protein Solubilization ................................................................................................ 37 2.17 Protein Recovery ....................................................................................................... 39 2.18 Functional Properties of Proteins in Foods ............................................................... 39 2.18.1 Solubility and absorption capacity ..................................................................... 40 2.18.2 Fat-binding and emulsification properties .......................................................... 41 2.18.3 Foaming properties ............................................................................................. 42 2.18.4 Gelation .............................................................................................................. 42 2.19 Effect of Drying Methods on the Functional Properties of Protein Isolate ............... 43 2.20 Protein Separation ..................................................................................................... 44 CHAPTER THREE ............................................................................................................... 45 3.0 MATERIALS AND METHODS .................................................................................... 45 3.1 Materials ...................................................................................................................... 45 3.1.1 Source of raw materials ........................................................................................ 45 3.1.2 Sample preparation ............................................................................................... 45 3.2 Methods ....................................................................................................................... 46 3.2.1 Proximate analyses ............................................................................................... 46 3.2.2 Extraction design of protein extract ..................................................................... 49 3.2.3 Extraction of protein ............................................................................................. 52 3.2.4 Protein