(558c) Two-Phase Kinetics of Enzymatic Depolymerization of Hemicellulose to Soluble Sugars for Liquid Biofuel Production

Hemicellulose is the second most abundant polysaccharide in nature, available not only in its pure form but also as a constituent of all lignocelluloses. The conversion of hemicelluoses to soluble sugars in parallel to its cellulosic counterpart would make liquid biofuels competitive in the market. Of the three key conversion steps (pretreatment of biomass, enzymatic hydrolysis of pretreated biomass to soluble sugars, ethanol from sugars by fermentation), hydrolysis is the slowest and the rate limiting step [1]. Adsorption of the liquid phase enzymes (endo- and exo-xylanases) to the insoluble solid substrate (xylan) is the first step in hemicellulose hydrolysis, followed by enzyme-catalyzed xylan depolymerization reactions in the solid and liquid phases. This work uses a coupled theoretical and experimental framework [2] to quantify the kinetics of product-inhibited solid and liquid phase depolymerization reactions that produce xylose and other reducing (soluble) sugars. We experimentally determine the type of adsorption of the enzyme on the solid substrate, the type of product inhibition in the solid and liquid phases, the transition of two-phase hydrolysis to single in batch reactors. Model-experiment comparisons allow us to quantify the xylobiose inhibition constants in the two phases. Our experiments show that the hydrolysis system transitions from a two-phase (solid-liquid) to a single (liquid) phase process at 11 hours. The enzymatic hydrolysis in both liquid and solid phases is found to be non-competitively inhibited by the product (xylose), with the inhibition being more potent in the liquid phase. Our multi-step reaction kinetic model is simulated using the liquid and solid phase kinetic data and validated with our experimentally measured temporal dynamics of xylose and reducing sugar yields in the liquid phase, and of the solid phase hemicellulose concentrations. We show that the product inhibition in the solid phase increases with the substrate loading, thus lowering the yield of reducing sugar, while the liquid phase product inhibition decreases with substrate loading, thus increasing the xylose yield. Our substrate inhibition studies allow us to recommend 5 mg/ml as the optimum xylan loading, above which substrate inhibition is observed to set in.

References:

[1] Chakraborty, S., Raju, S., and Pal, R., "A multiscale three-zone reactive mixing model for engineering a scale separation in enzymatic hydrolysis of cellulose," Bioresource Technology, 173, pp 140-147 (2014)

[2] Dutta, S.K., and Chakraborty, S., â??â??Kinetic analysis of two-phase enzymatic hydrolysis of hemicellulose of xylan type,â??â?? Bioresource Technology, 198, pp 642-650 (2015)