Direct quantitative determination of adsorbed cellulase on lignocellulosic biomass with its application to study cellulase desorption for potential recycling

Effective hydrolysis of pretreated lignocellulose mediated by cellulase requires an in-depth understanding of cellulase adsorption and desorption. Here we developed a simple method for determining the adsorbed cellulase on cellulosic materials or pretreated lignocellulose, which involves (i) hydrolysis of adsorbed cellulase in the presence of 10 M of NaOH at 121 °C for 20 min, and (ii) the ninhydrin assay for the amino acids released from the hydrolyzed cellulase. The major lignocellulosic components (i.e., cellulose, hemicellulose, and lignin) did not interfere with the ninhydrin assay. A number of cellulase desorption methods were investigated: pH change as well as the use of detergents, high salt solution, and polyhydric alcohols. The pH adjustment to 13.0 and the elution by 72% ethylene glycol at neutral pH were among the most efficient approaches for desorbing the adsorbed cellulase. For the recycling of active cellulase, a modest pH adjustment to 10.0 may be a low-cost viable method to desorb active cellulase. It was found that more than 90% of cellulase for hydrolysis of the pretreated corn stover could be recycled by washing at pH 10.0.