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TITLE
CERTIFICATE 1
CERTIFICATE-2
DECLARATION
ACKNOWLEDGEMENT
CONTENTS
ABBREVIATIONS AND NOTATIONS
PREFACE
I. INTRODUCTION
II. REVIEW OF LITERATURE
2.1 Cellulose
2.2 Cellulase
2.3 Isolation of cellulolytic organisms from different sources
2.4 Cellulase production
2.5 Kinetics of cellulase production
2.6 Pretreatment
2.7 Fungal protein production
2.8 Purification of the enzyme
2.9 Saccharification of cellulose wastes
2.10 Kinetics of the enzymatic hydrolysis of cellulose
2.11 Alcohol production
2.12 Oyster mushroom
III. MATERIALS AND METHODS
3.1 Cellulase producing bacteria and fungi fromnatural environment
3.1.1 Preparation of media
3.1.1.1 Modified Hans agar medium (pH 6.5)
3.1.1.2 Modified Czapeks agar medium (pH 6.4-7.0)
3.1.1.2 Mandels and Reese agar medium (pH 5.3)
3.1.1.3 Nutrient agar medium (pH 6.2)
3.1.1.4 Potato dextrose agar medium (pR 6.0-6.5)
3.1.2 Enumeration of microbial population and isolationof cellulolytic bacteria and fungi
3.1.2.1 Isolation and maintenance of cultures
3.1.2.2 Identification of bacteria and fungi
3.1.3 Screening of cellulolytic microorganisms
3.1.3.1 Primary screening
3.1.3.2 Secondary screening
3.1.3.2.1 Media
3.1.3.2.2. Inoculation procedure for bacteria
3.1.3.2.3 Inoculum procedure for fungi
3.1.3.2.4 Measurement of growth
3.1.3.2.5 Enzyme production
3.1.4 Assay of cellulase
3.1.4.1 Assay of C -cellulase
3.1.4.2 Assay of C1-cellulase
3.1.4.3 Combined assay of C1 and Cx-cellulase
3.1.4.4 Assay of fi-glucosidase
3.1.5 Saccharification of cellulose wastes
3.1.5.1 Estimation of total sugars
3.1.5.2 Estimation of glucose
3.2 Screening of ligninolytic organisms
3.2.1 Crawfords agar medium (pH 7.0)
3.3 Growth studies
3.3.1 Effect of physico-chemical factors on growth andenzyme production
3.3.2 Effect of agitation
3.3.3 Effect of carbon sources
3.3.4 Effect of nitrogen sources
3.4 Growth curve
3.5 Composition and pretreatment studies ofcellulose waste
3.5.1 Samples and sampling
3.5.2 Composition of the sample
3.5.2.1 Estimation of cellulose
3.5.2.2 Estimation of hemicellulose
3.5.2.3 Estimation of starch
3.5.2.4 Estimation of pectin
3.5.2.5 Estimation of lignin
3.5.2.6 Estimation of protein
3.5.2.7 Estimation of total lipids
3.6 Pretreatment studies of cellulose waste
3.6.1 Physical pretreatment
3.6.2 Chemical pretreatment
3.6.2.1 Sodium hydroxide pretreatment
3.6.2.2 Sodium hydroxide-acetic acid pretreatment
3.6.2.3 Chloroform pretreatment
3.6.2.4 Acid pretreatment
3.6.2.5 Sodium sulphite pretreatment
3.6.2.6 Peracetic acid pretreatment
3.6.2.7 Butanol pretreatment
3.6.2.8 Hydrogen peroxide-ferrous salt pretreatment
3.6.2.9 Hydrogen peroxide-manganous salt pretreatment
3.6.2.10 Hydrochloric acid-zinc chloride pretreatment
3.6.2.11 Acetic acid-hydrogen peroxide pretreatment
3.7 Enzyme Studies
3.7.1 Isolation of enzyme
3.7.1.1 Media
3.7.1.2 Enzyme production in the medium
3.7.2 Purification of enzyme
3.7.2.1 Ammonium sulphate precipitation
3.7.2.2 Desalting of the enzyme
3.7.2.3 Fractionation of the enzyme
3.8 Characterization of cellulase
3.9 Bioconversion of tapioca waste and water hyacinth
3.9.1 Solid state fermentation
3.9.1.1 Cellulose source
3.9.1.2 Media
3.9.1.3 Microorganisms
3.9.1.4 Inoculum preparation
3.9.1.5 Inoculation procedure
3.9.1.6 Fermentation
3.9.2 Conversion of cellulose wastes by enzyme
3.9.2.1 Production of cellulase
3.9.2.2 Samples
3.9.2.3 Saccharification
3.9.3 Saccharification of cellulose wastes using termitegut extract
3.9.3.1 Samples
3.9.3.2 Preparation of termite gut extract
3.9.3.3 Saccharification
3.10 Alcohol fermentation
3.10.1 Microorganism
3.10.1.1. Media
3.10.1.1.1 Glucose yeast extract agar medium (pH 7.0)
3.10.1.1.2 culture medium (pH 5.0)
3.10.2 Fermentation media
3.10.3 Fermentation of glucose to alcohol
3.10.4 Estimation of alcohol
3.10.5 Effect of pH
3.10.6 Effect of temperature
3.10.7 Effect of substrate concentration
3.10.8 Effect of incubation period
3.11 Oyster mushroom farming
3.11.1 Preparation of spawn
3.11.2 Substrate preparation
IV. RESULTS AND DISCUSSION
PART I SCREENING OF CELLULOLYTIC MICROORGANISMS
4.1 Isolation of cellulolytic bacteria and fungi
4.1.1 Primary screening and selection
4.1.2 Identification of selected strains
4.1.3 Secondary screening
4.2 Optimization of Hicrobial Growth andEnzyme Production
4.2.1 Effect of temperature
4.2.2 Effect of pH
4.2.3 Effect of substrate concentration
4.2.4 Effect of incubation period
4.2.5 Effect of agitation
4.2.6 Effect of carbon sources
4.2.7 Effect of nitrogen sources
4.3 Growth curve
PART II COMPOSITION AND PRETREATMENT STUDIES OF WASTE SAMPLES
4.4 Composition
4.5 Pretreatment Studies
4.5.1 Physical pretreatments
4.6 Chemical Pretreatments
4.6.1 Effect of sodium hydroxide
4.6.2 Effect of sodium hydroxide-acetic acid
4.6.3 Effect of chloroform
4.6.4 Effect of hydrochloric acid
4.6.5 Effect of sodium sulphite
4.6.6 Effect of peracetic acid
4.6.7 Effect of butanol
4.6.7 Effect of hydrogen peroxide and ferrous salt
4.6.8 Effect of hydrogen peroxide and manganous salt
4.6.9 Effect of hydrochloric acid and zinc chloride
4.6.10 Effect of acetic acid and hydrogen peroxide
PART III PURIFICATION AND CEARACTERIZATION OF ENZYME
4.7 Purification
4.8 Enzyme studies
4.8.1 Effect of pH on the activity of the enzyme
4.8.2 Effect of temperature on the activity of the enzyme
4.8.3 Effect of substrate concentration on the activityof the enzyme
4.8.4 Effect of incubation period on the activity ofthe enzyme
4.8.5 Effect of agitation on the activity of the enzyme
PART IV BIOCONVERSION OF TAPIOCA WASTE AND WATER HYACINTH
4.9 Bioconversion
4.9.1 Solid state fermentation by the action ofmicroorganisms
4.9.2 Action of cellulolytic microbesand ligninolytic fungi
4.9.3 Bioconversion of cellulose waste by enzyme
4.9.4 Bioconversion of cellulose waste using termite gutextract
PART V ALCOHOL FERMENTATION
4.10 Fermentation Studies
4.10.1 Effect of pH on alcohol fermentation
4.10.2 Effect of temperature
4.10.3 Effect of incubation period
4.10.4 Effect of substrate concentration
4.10.5 Alcohol fennentati.on of the fermentable sugars
4.10.6 Alcohol fermentation of the fermentable sugars
PART VI FARMING OF OYSTER MUSHROOM
4.11 Farming of oyster mushroom
Plate 1a -P. florida mycelial growth on tapioca waste.
Plate 1b Oyster mushroom (P. florida) on tapioca waste.
Plate 2a florida mycellar growth on water hyacinth.
Plate 2b Oyster mushroom (p.florida) on water hyacith.
V. SUMMARY
REFERENCES