The relation between the supramolecular structure of cellulose and its hydrolysability

Doctoral thesis, 2015

The liberation of fermentable sugars from cellulosic biomass during enzymatic hydrolysis is often incomplete. One of the factors limiting the efficiency of enzymatic hydrolysis is the structural properties of cellulose. The aim of the work presented in this thesis was to increase our understanding of the relation between enzymatic hydrolysis and the structural properties of cellulosic substrates. The enzymatic hydrolysis of a number of cellulosic substrates derived from softwood preparations used in the pulp and paper industry, as well as model substrates, were studied. The differences in cellulosic substrates before and after enzymatic hydrolysis are described on the nanometre scale in terms of their supramolecular structure, i.e. the lateral dimensions of fibrils and fibril aggregates, the accessible surface area, the crystallinity and porosity, using solid-state nuclear magnetic resonance spectroscopy. The substrates were imaged and structural changes in the cellulosic substrates were characterized in real time on the micrometre scale in terms of their molecular density, ordering and autofluorescence, employing nonlinear optical microscopy. A strong correlation was found between the average pore size and the specific surface area of the starting material and the enzymatic conversion yield. The overall degree of crystallinity and the lateral dimensions of the fibrils increased in some samples as a result of hydrolysis. Avicel had a higher carbon–hydrogen bond density and a different pattern of ordered structures than the never-dried pulp fibres, possibly reflecting the collapse of the macromolecular structures during drying and rewetting. Monitoring of the substrates during enzymatic hydrolysis revealed substrate-characteristic hydrolysis pattern. The response of the most widely studied filamentous fungus for cellulase production, Trichoderma reesei, to cellulosic substrates with different supramolecular structures was studied. Substantial differences were found in the profile of the enzymes produced, despite the fact that there were only minor differences in the chemical composition of the cellulose-rich substrates. Culture filtrates from five filamentous fungi cultivations were evaluated regarding their ability to improve saccharification of the industrial cellulase cocktail Celluclast 1.5L. It was demonstrated that supplementing commercial cocktails with enzymes from carefully selected fungi can result in significantly more efficient saccharification of biomass.