Production of fuel ethanol and other chemicals from renewable biomass is of strategic importance.  To effectively convert the carbohydrates in complex and recalcitrant biomass, mechanical or chemical pretreatments are often required proceeding enzymatic hydrolysis and microbial fermentation.  However, pretreatments may negatively affect the enzymatic hydrolysis by releasing by-products inhibitory to cellulases, or modifying cellulose by oxidation or other processes.  To study how oxidative modification may impact cellulose’s reactivity toward the hydrolysis by cellulases, we prepared three cellulose substrates by cupric ion and hypochlorite oxidations, and subjected the derived celluloses to the hydrolysis of several Cel7 and Cel6 cellobiohydrolase homologues, two GH61 cellulase enhancing proteins, and one cellulolytic Trichoderma reesei extracellular enzyme mixture.  We observed a profound decrease of enzymatic hydrolysis on the oxidized celluloses.  The effect was attributed to the interference, from oxidized functional groups in cellulose, on its binding/activation in the active pocket/tunnel of cellobiohydrolases.  Potential implication of the observed effect from cellulose oxidation on pretreatment optimization was discussed.