(490ac) Enzymatic Hydrolysis of Cellulosic Substrates Under High Solid Loading

Enzymatic hydrolysis of cellulosic substrate is affected by many parameters. One of the unexplained behaviors with regard to cellulase reaction is that the glucan digestibility normally decreases under high solid loadings. A comprehensive experimental study was conducted to verify this point using Spezyme CP and Avicel. To this end, the initial hydrolytic reaction rates the overall yields were measure at various substrate concentrations while keeping the substrate-enzyme ratio constant. The results have shown that increasing substrate concentration led to decreased reaction rate and yield despite that the enzyme loading per gram glucan was fixed. The decrease of yield was much more than that expected from glucose inhibition. The enzyme activity was then measured taking time-course samples. The method of activity measurement involved desorption of enzymes from the substrate to account for all the enzymes existing in the reactor. The measured specific activity of cellulose enzyme (FPU/mg) did not change during the course of the reaction for all substrate loadings applied. Upon this finding, initial reaction experiments were pursued using Avicel as the substrate. Lineweaver-Burk analysis of the data indicated that the Km value of the cellulose measured against Avicel increased proportionally with the substrate concentration over the range of 1-5% w/v. This indicates that the affinity of enzyme with substrate decreases at high solid loading, which led to decreased sugar yields at high solid loading. To further verify this point enzyme adsorption kinetics and isotherms were investigated. The results are analyzed in connection with variation of binding capacity of enzymes to substrate and change of synergism among the different cellulose components.