Date of Graduation

12-2015

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Engineering (PhD)

Degree Level

Graduate

Department

Biological and Agricultural Engineering

Advisor

Danielle Julie Carrier

Committee Member

Robert Beitle

Second Committee Member

Edgar C. Clausen

Third Committee Member

Thomas A. Costello

Keywords

Applied sciences; Antimicrobial activity; Antioxidant activity; Bioenergy; Cellulase inhibition; Dilute acid pretreatment; Phytochemical extraction

Abstract

The effect of compounds produced during biomass pretreatment on cellulolytic enzyme was investigated. Liquid prehydrolyzates were prepared by pretreating switchgrass using 24 combinations of temperature, time, and sulfuric acid concentration based on a full factorial design. Temperature was varied from 140°C to 180°C; time ranged from 10 to 40 min; and the sulfuric acid concentrations were 0.5% or 1% (v/v). Identified products in the prehydrolyzates included xylose, glucose, hydroxymethylfurfural (HMF), furfural, acetic acid, formic acid, and phenolic compounds at concentration ranging from 0 to 21.4 g/L. Pretreatment conditions significantly affected the concentrations of compounds detected in prehydrolyzates. When assayed in the presence of switchgrass prehydrolyzates against model substrates, activities of cellulase, betaglucosidase, and exoglucanase, were significantly reduced by at least 16%, 31.8%, and 57.8%, respectively, as compared to the control. A strong positive correlation between inhibition of betaglucosidase and concentration of glucose, acetic acid, and furans in prehydrolyzate was established. Exoglucanase inhibition correlated with the presence of phenolic compounds and acetic acid. The prehydrolyzate, prepared at 160°C, 30 min, and 1% acid, was fractionated by centrifugal partition chromatography (CPC) into six fractions; the inhibition effect of these fractions on betaglucosidase and exoglucanase was determined. The initial hydrolysis rate of cellobiose by betaglucosidase was significantly reduced by the CPC sugar-rich fraction; however, exoglucanase was deactivated by the CPC phenolic-rich fraction. Finally, biological activities of water-extracted compounds from sweetgum bark and their effect on cellulase was investigated. It was determined that 12% of solid content of the bark extract could be accounted by phenolic compounds with gallic acid identified as the most concentrated phytochemical. Sweetgum bark extract inhibited Staphylococcus aureus growth and copper-induced peroxidation of human low-density lipoprotein, confirming antimicrobial and antioxidant activities of the extract. On the other hand, bark extract inhibited cellulase cocktail activity by reducing cellulose hydrolysis by 82.32% after 48 h of incubation. Overall, phenolic compounds generated from biomass fractionation are important players in cellulolytic enzyme inhibition; removal of biomass extractives prior to pretreatment could reduce inhibitory compounds in prehydrolyzate while generating phytochemicals with societal benefits.

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