Substrate Characteristics That Influence the Filter Paper Assay’s Ability to Predict the Hydrolytic Potential of Cellulase Mixtures

Abstract

Currently, filter paper assay (FPA) is used to measure the hydrolytic potential of cellulase enzyme mixtures that are used to hydrolyze a range of cellulosic substrates. However, some of the Whatman No. 1 filter paper substrate characteristics, such as its high cellulose content, cellulose accessibility, dried nature, etc., compromise the ability of the FPA to predict how well a complex cellulase cocktail will hydrolyze pretreated lignocellulosic substrates. The influence of factors such as drying, papermaking, and the presence of lignin and hemicellulose on the FPA was assessed by producing and using “model/paper sheets” as a substitute for the Whatman No. 1 filter paper. Drying was shown to have a major, negative influence resulting in a 60% reduction in the assessed cellulase activity. Similarly, a 53% decrease in cellulase activity resulted when an organosolv-pretreated hardwood containing lignin and hemicellulose was used instead of the Whatman No. 1 filter paper. It was apparent that the FPA was a poor predictor of the hydrolytic potential of an enzyme mixture on cellulosic/lignocellulosic substrates. However, other than assessing the ease of hydrolysis of a specific substrate under a defined set of conditions, it is likely that the FPA will continue to be used to indicate the broad, hydrolytic potential of an enzyme mixture in terms of the amount of protein required to hydrolyze the cellulosic component over a defined set of temperature, time, and pH conditions.