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Type of Document Dissertation Author Sun, Ye , URN etd-08152002-220933 Title Enzymatic Hydrolysis of Rye Straw and Bermudagrass for Ethanol Production Degree PhD Graduate Program Biological and Agricultural Engineering Advisory Committee
Advisor Name Title Philip Westerman Committee Chair Jiayang Cheng Committee Co-Chair Joel Ducoste Committee Member John Classen Committee Member Yuri Yamamoto Committee Member Keywords
- transgenic duckweed and cellulases
- endoglucanase E1
Date of Defense 2002-08-12 Availability unrestricted Abstract Dilute sulfuric acid pretreatment of rye straw and bermudagrass was investigatedunder acid concentrations of 0.6% to 1.5% (w/w) and residence time 30 min to 90 min.
The pretreatment effectively solubilized the hemicellulose components to monomeric
sugars such as xylose, arabinose, and galactose. Cellulose in the rye straw was not
hydrolyzed into glucose during the acid pretreatment, while part of the cellulose in the
bermudagrass was converted into glucose by the acid pretreatment. After enzymatic
hydrolysis with excessive cellulases, the glucose yields of 30% to 52% and 46% to 81%
of the theoretical potentials were obtained for rye straw and bermudagrass, respectively.
The pretreated solid residues were hydrolyzed with cellulases supplemented with
β-glucosidase. The addition of β-glucosidase greatly improved the glucose production
rate. There was no cellobiose accumulation when the β-glucosidase loading was up to 25
CBU/g (CBU, cellobiase unit, expressed as µmol of cellobiose that is converted into
glucose per minute). The conversion rate was 45% for bermudagrass hydrolyzed with
cellulases of 10 FPU/g (FPU, filter paper unit, expressed as µmol of glucose produced
per min with filter paper as a substrate) and β-glucosidase of 25 CBU/g. The further
increase of enzyme loadings did not significantly improve the glucose yield. Rye straw
was more resistant to enzymatic hydrolysis than bermudagrass. The conversion rate was
38% with the additions of cellulases at 15 FPU/g and β-glucosidase at 25 CBU/g rye
straw.
The production of cellulase enzymes in transgenic plants has the potential to
significantly reduce enzyme costs. Expression of cellulase enzyme, Acidothermus
cellulolyticus E1 endoglucanase, in transgenic duckweed Lemna minor was examined.
The recombinant E1 protein was biologically active with the expression level of 0.24% of
total soluble protein. HEPES (N-[2-Hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid])
buffer (pH = 8) extracted more proteins including E1 from duckweed fronds than sodium
citrate buffer (pH = 4.8) and sodium acetate buffer (pH = 5). The E1 protein was
thermotolerant and exhibited the optimal enzyme activity at temperature 80°C and pH 5.
The E1 activity remained unchanged after heating at 60°C for 6 h. However, it was
inactivated after 15-min heating at 90°C.
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