This study quantified the activities of enzymes in extracellular polymeric substances ( EPS) and in pellets. Seven commonly adopted extraction schemes were utilized to extract from aerobic flocs the contained EPS, which were further categorized into loosely bound ( LB) and tightly bound ( TB) fractions. Ultrasonication effectively extracted the EPS from sludge flocs. Enzyme assay tests showed that the protease activity was localized mainly on the pellets, amylase and alpha-glucosidase activities were largely bound with LB-EPS, and few protease, alpha-amylase, or alpha-glucosidase activities were associated with the TB-EPS fraction. There exists no correlation between the biochemical compositions of EPS and the distribution of enzyme activities in the sludge matrix. The 44-65% of alpha-amylase and 59-100% of alpha-glucosidase activities noted with the LB-EPS indicate heterogeneous hydrolysis patterns in the sludge flocs with proteins and carbohydrates.