Enzymatic Hydrolysis of Biomass: Effects of Crystallinity, Particle Size, and Lignin Removal

Enzymatic hydrolysis of cellulose in plant and wood cell walls is expected to be affected by its chemical composition as well as structural and morphological features. In the present study, different crystallinity cellulose samples and varying size fractionated loblolly pine wood cell wall particles were hydrolyzed using a mixture of Celluclast and Novozyme 188. In pure cellulose samples of Whatman CC31, at lower crystallinity, higher hydrolysis rates and higher conversion to glucose were observed. Percent cellulose hydrolysis was linearly but inversely correlated with percent crystallinity. After 72 h, the extent of hydrolysis for highly crystalline and completely amorphous cellulose samples was 40% and 90%, respectively. In the case of loblolly pine, softwood, four different size Wiley-milled wood fractions along with a ball milled sample were hydrolyzed. Although, for the four of the five fractions, where the cellulose crystallinity was similar, there was some increase in enzyme hydrolysis with reduced particle size, most of the increased hydrolysis occurred for the ball milled wood indicating that the destruction of crystallinity and accompanying changes had a major impact. Moreover, to examine the impact of lignin removal, partially delignified pine samples of the four fractions and increasingly delignified samples of a chosen loblolly pine fraction were subjected to enzymatic saccharification. In the former case, lignin removal was found to be associated with significantly increased hydrolysis extent of hydrolysis at 72 h for the four wood fractions increased on average by 10 fold. Results of the experiments on the increasingly delignified samples indicated higher saccharification with increased lignin removal. Lastly, when the hydrolysis data between the similar crystallinity cellulose and wood cell wall were compared, at 72 h, the cell wall conversion was found to be 4 times slower. These results taken together highlighted that in the biomass hydrolysis cellulose crystallinity and cell wall composition are not as important as the ultrastructural changes that ensued upon acid chlorite delignification.