1
GLUCOSE PRODUCTION FROM CASSAVA STARCH BY MONOCULTURE
AND COCULTURE OF RAGI TAPAI AND SACCHAROMYCES CEREVISIAE
MA SIEW VOON
A thesis submitted in partial fulfillment of the
requirements for the award of the degree of
Bachelor of Engineering (Chemical)
Faculty of Chemical Engineering
Universiti Teknologi Malaysia, Johor
JANUARY 2013
v
ABSTRACT
Bioethanol is in the upsurge trend as the potential alternative fuel for gasoline.
Bioethanol is produced from fermentation of reducing sugar. Production of sugar as
feedstocks for bioethanol is the core of this research. The first objectives of this
research is to evaluate the overall trend in terms of glucose production via
simultaneous saccharification and fermentation of cassava starch in room
temperature by using indigenous culture called “ragi tapai”, S. cerevisae and a
coculture of “ragi tapai” and S.cerevisiae. Secondly, the effects of various parameters
such as temperature, culture time and starch to inoculum ratio are investigated on
hydrolysis of starch by using “ragi tapai” only. Agitation speed, inoculum size and
nitrogen source are fixed during the experiment. The experiment is conducted for
72hours and glucose yield is measured at different time interval. The range of the
factors investigated is room temperature and 35°C (temperature), 1 hour and 3 hours
(culture time), 5w/v%, 10w/v% and 15w/v% (starch concentration). The yield of
glucose is analyzed by using DNS colorimetric method. Based on the findings
obtained from the research, “ragi tapai” showed the highest potential in hydrolyzing
polysaccharides of starch molecule into monosaccharide glucose in room
temperature. S.cerevisiae showed the lowest hydrolyzing ability while coculture of
both “ragi tapai” and S.cerevisiae showed fluctuation in glucose production. “Ragi
tapai” has showed the best performance in these conditions; of room temperature, 1
hour cultured and 1:1 starch to inoculums ratio. The highest glucose generated during
the study is 15.8g/L and this yield is 31.6% with respect to dry weight basis of
cassava starch powder used and 11.3% with respect to wet weight basis.
vi
ABSTRAK
Bioethanol adalah dalam trend peningkatan sebagai bahan api alternatif yang
berpotensi untuk petrol. Bioetanol dihasilkan daripada penapaian gula ringkas.
Penghasilan gula sebagai bahan mentah untuk bioethanol adalah teras penyelidikan
ini. Objektif pertama kajian ini adalah untuk menilai trend keseluruhan dari segi
penghasilan glukosa melalui saccharification serentak dan penapaian kanji ubi kayu
pada suhu bilik dengan menggunakan monokultur dan cokultur komersial ragi tapai
dan S.cerevisiae. Kedua, kesan pelbagai parameter seperti suhu, kultur masa dan
nisbah kanji kepada inokulum disiasat pada hidrolisis kanji oleh monokultur ragi
tapai sahaja. Kelajuan putaran, saiz inokulum dan sumber nitrogen ditetapkan semasa
eksperimen. Eksperimen dijalankan untuk 72hours dan hasil glukosa diukur untuk
selang masa yang berbeza. Pelbagai faktor yang dikaji adalah suhu bilik dan 35 °C
(suhu), 1 jam dan 3 jam (kultur masa), 5w/v%, 10w/v% dan 15w/v% (kanji
kepekatan). Hasil glukosa dianalisis dengan menggunakan kaedah kolorimetrik DNS.
Berdasarkan hasil kajian, monokultur ragi tapai mempunyai potensi tertinggi dalam
menukarkan polisakarida molekul kanji menjadi glukosa monosakarida pada suhu
bilik. S.cerevisiae menunjukkan keupayaan terendah dalam penghasilan glukosa
manakala cokultur kedua-dua yis menunjukkan turun naik dalam pengeluaran
glukosa. Monokultur ragi tapai telah menunjukkan prestasi yang terbaik dalam
keadaan suhu bilik, 1 jam pengulturan dan nisbah 1:1 kanji kepada inokulum.
Glukosa tertinggi yang dihasilkan semasa penyelidikan adalah 15.8g / L dan hasil ini
adalah 31.6% dengan asas berat kering serbuk kanji ubi kayu digunakan dan 11.3%
dengan asas berat basah kanji.
vii
TABLE OF CONTENTS
CHAPTER
1
2
TITLE
PAGE
DECLARATION
ii
DEDICATION
iii
ACKNOWLEDGEMENT
iv
ABSTRACT
v
ABSTRAK
vi
TABLE OF CONTENTS
vii
LIST OF TABLES
x
LIST OF FIGURES
xi
LIST OF ABBREVIATIONS
xiii
INTRODUCTION
1
1.1
Background of Study
1
1.2
Problem Statement
2
1.3
Objective of Study
3
1.4
Scope of Study
4
1.5
Significance of Study
5
LITERATURE REVIEW
6
2.1
Biofuel
6
2.2
Bioethanol
7
2.3
Bioethanol Economics in Malaysia
8
2.4
Raw Material for Bioethanol Production
9
viii
2.5
Cassava
11
2.6
Starch
13
2.7
Glucose
14
2.8
Yeast
15
2.8.1 Ragi tapai
16
2.8.2 Saccharomyces cerevisiae
18
2.9
Microbial Culture
19
2.10
Pretreatment/ Gelatinization Process
20
2.11
Liquefaction and Saccharification Process
20
2.12
Ethanol Fermentation
21
2.13
Parameters Affect Hydrolysis Rate
22
2.13.1 Temperature
22
2.13.2 Culture Time
23
2.13.3 Substrate Concentration
24
DNS Glucose Analysis Method
25
2.14
3
METHODOLOGY
26
3.1
Material and Reagents
26
3.2
Experimental Equipment and Apparatus
27
3.3
Experimental Procedure
27
3.3.1 Sample Preparation
28
3.3.2 Yeast and Culture Medium
28
3.3.3 Glucose Production by Monoculture and
28
Coculture of Yeast
3.3.4 Evaluation for Sugar Hydrolysis Performance
29
3.3.4.1 Effect of Cassava Starch to Inoculum Ratio 30
3.4
3.3.4.2 Effect of Culture Time
30
3.3.4.3 Effect of Temperature
31
Analysis Method
31
3.4.1 Glucose Standard Preparation
31
3.4.2 Determination of Glucose by DNS Method
32
ix
4
RESULTS AND DISCUSSIONS
34
4.1
Standard Calibration Curve for Glucose
34
4.2
Sugar Production using Monoculture Ragi Tapai
36
4.3
Sugar Production using Monoculture S. cerevisiae
38
4.4
Sugar Production using Coculture Ragi Tapai and
39
S.cerevisiae
4.5
5
Saccharification of Starch by Ragi tapai
41
4.5.1 Effect of Cassava Starch to Inoculum Ratio
41
4.5.2 Effect of Culture Time
43
4.5.3 Effect of Temperature
44
CONCLUSIONS AND RECOMMENDATIONS
46
5.1
Conclusions
46
5.2
Recommendations
47
REFERENCES
49
APPENDIX
56
49
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