Deactivation of cellulases by phenols

Pretreatment of lignocellulosic materials may result in the release of inhibitors and deactivators of cellulose enzyme hydrolysis. We report the identification of phenols with major inhibition and/or deactivation effect on enzymes used for conversion of cellulose to ethanol. The inhibition effects were measured by combining the inhibitors (phenols) with enzyme and substrate immediately at the beginning of the assay. The deactivation effects were determined by pre-incubating phenols with cellulases or β-glucosidases for specified periods of time, prior to the respective enzyme assays. Tannic, gallic, hydroxy-cinnamic, and 4-hydroxybenzoic acids, together with vanillin caused 20-80% deactivation of cellulases and/or β-glucosidases after 24h of pre-incubation while enzymes pre-incubated in buffer alone retained all of their activity. The strength of the inhibition or deactivation effect depended on the type of enzyme, the microorganism from which the enzyme was derived, and the type of phenolic compounds present. β-Glucosidase from Aspergillus niger was the most resistant to inhibition and deactivation, requiring about 5 and 10-fold higher concentrations, respectively, for the same levels of inhibition or deactivation as observed for enzymes from Trichoderma reesei. Of the phenol molecules tested, tannic acid was the single, most damaging aromatic compound that caused both deactivation and reversible loss (inhibition) of all of enzyme activities tested.