Pre Treatment With White Rot Fungi For Ethanol Biology Essay

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In the present study, Rubber wood was used as the raw material for bioethanol production. The goal of this study was to investigate the efficiency of biological pretreatment with Ceriporiopsis subvermispora ATCC 90467, Trametes versicolor ATCC 20869, and combination of both fungi for conversion of Rubber wood to ethanol. Change in chemical composition, structural modification, and their susceptibility to enzymatic saccharification and ethanol production in the degraded wood were analyzed. Result of this study showed that selective lignin-degrading fungus C. subvermispora had greater selectivity for lignin degradation. After 90 days of pre-treatment with C. subvermispora, lignin and hemicellulose loss were the highest at 45.06 % and 42.08 %, respectively among the tested samples. While, cellulose loss was lower at 9.50 % compared to those of T. versicolor and combine over a period of 90 days. The influence of particle size (0.250, 0.5, and 1 mm) on pretreatment effectiveness by C. subvermispora also was examined. The Rubber wood with the particle size of 1mm was efficiently degraded compared with smaller particle size. To evaluate the biological pre-treatment, cellulose in the pretreated woods was hydrolyzed using Celluclast 1.5 L and Novozyme 188 at 50°C for 168 hours and the released sugars were converted to ethanol by simultaneous saccharification and fermentation process (SSF) using yeast Saccharomyces cerevisiae D5A at 37 °C for 120 hours. A study on hydrolysis of Rubber wood treated with C. subvermispora, T. versicolor, and combine for 90 days resulted in an increase sugar yield about 27.67%, 16.23%, and 14.20%, respectively compared with untreated Rubber wood (2.88 %). The sample obtained using the best pretreatment (sample pretreated by C. subvermispora) was used for bioethanol production. After 120 hours, the maximum ethanol concentration and yield were obtained from fungal pretreated samples for 90 days of cultivation time (17.9 g/L and 53 % of theoretical maximum, respectively). The results obtained demonstrate that white rot fungus C. subevermispora provides an effective method for improving the enzymatic hydrolysis and ethanol production of Rubber wood. The results also demonstrate that Rubber wood is a potential raw material for bioethanol production.

TABLE OF CONTENTS

Page

DEDICATION

ABSTRACT

ABSTRAK

ACKNOWLEDGMENTS

APPROVAL

DECLARATION

LIST OF TABLES

LIST OF FIGURES

LIST OF ABREVIATIONS

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iii

vi

ix

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xvi

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CHAPTER

INTRODUCTION

1.1 General background

Objective

2 LITERATURE REVIEW

2.1 General background

2.2 Classification of bioethanol production

2.2.1 First generation bioethanol

2.2.2.1 Limitations of first generation bioethanol

2.2.1 Second generation bioethanol

2.3 Overview biomass in Malaysia

2.4 Why choose Rubber wood?

2.5 Lignocellulosic biomass

2.5.1 Components of lignocellulosic materials

2.5.1.1 Extraneous substances

2.5.1.2 Polysaccharides

2.5.1.2.1 Cellulose

2.5.1.2.2 Hemicellulose

2.5.1.2.3 Lignin

2.5.2 Structure of lignocellulosic materials

2.6 Chalenges in bioethanol production from lignocelluloses

2.7 Pre-treatment

2.7.1 Physical pre-treatment

2.7.2 Physicochemical pre-treatment

2.7.3 Chemical pre-treatment

2.7.4 Biological pre-treatment

2.7.4.1 Microbial strains

2.7.4.1.1 Ceriporiopsis subvermispora

2.7.4.1.2 Trametes versicolor

2.8 Hydrolysis of cellulose

2.8.1 Acid hydrolysis

2.8.2 Enzymatic hydrolysis

2.8.2.1 Cellulase

2.8.2.1.1 Mechanism of cellulose

hydrolysis

2.8.3 End product inhibition

2.9 Fermentation

2.9.1 Separate hydrolysis and fermentation (SHF)

2.9.2 Simultaneous saccharification and fermentation (SSF)

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MATERIALS AND METHODS

Materials and reagents

3.1.1 Rubber wood

3.1.2 Chemical material

3.1.3 DNS reagent

3.1.4 Citrate buffer (0.05 M, pH = 4.8)

3.1.5 Enzyme

3.1.6 Media

3.1.7 Microorganisms

3.2 Preparation of Rubber wood

3.3 Characterization of Rubber wood

3.3.1 Chemical analysis

3.3.1.1 Determination of total solids

3.3.1.2 Determination of extractives

3.3.1.3 Determination of klason lignin

3.3.1.4 Determination of holocellulose

3.3.1.5 Determination of α-cellulose and

Hemicellulose

3.3.2 Fourier Transform Infrared Spectroscopy

(FTIR)

3.3.3 X-ray diffraction

3.3.4 Scanning electron microscopy (SEM)

3.4 Biological pre-treatment

3.4.1 Strain and inoculation

3.4.2 Solid state fermentation (SSF)

3.5 Enzyme hydrolysis

3.5.1 Determination of reducing Sugar by dinitrosalicylic

acid (DNS) method

3.6 Production of ethanol

3.7 Gas chromatography (GC)

3.8 Experimental design and statistical analysis

RESULT AND DISSCUSSION

Cmposition of Rubber wood

Effect of fungal pre-treatment on chemical composition

X-ray diffractions

FTIR analysis

Undecayed Rubber wood (control)

Wood decayed by C. subvermispora

Wood decayed by T. versicolor

Wood decayed by combination of both fungi

Comparison of Rubber wood decayed by C. subvermispora, T. versicolor, and combine

Effect of particle size on degradation of Rubber wood by C.

Subvermispora

Chemical composition analysis

X-ray diffraction

4.6 Scanning electron microscopy (SEM)

4.7 Enzymatic hydrolysis

4.7.1 Effect of pre-treatment time on enzymatic hydrolysis

4.7.2 Relationship between crystallinity index (CrI) and

reducing sugar yield

4.7.3 Relationship between lignin content and reducing

sugar yield

4.8 Production of bioethanol

SUMMARY, CONCOLUSION, AND RECOMANDATIONS

FOR FUTURE RESEARCH

Summary

Conclusion

Recommendations for future research