The effects of heat pretreatment on waste sludge hydrolysis were investigated in this study. Heat pretreatment was conducted at 65 degrees C, 80 degrees C, 100 degrees C and 121 degrees C for 5 min, 10 min, 15 min, 20 min, 25 min and 30 min. Not only analyzed the changes of SCOD (Soluble chemical oxygen demand), carbohydrate and protein, but also evaluated the structural and functional properties of organics in extracellular polymeric substances (EPS) and dissolved organic matters (DOM) by using three-dimensional excitationemission matrix (3D-EEM) fluorescence spectroscopy with fluorescence regional integration (FRI) analysis. The SCOD in DOM increased with pretreated temperatures. The optimal heat hydrolysis temperature and time were selected by further studying the biodegradable and non-biodegradable components. After treated at 80 degrees C for 25 min, the fluorescence intensity and percent fluorescence response (P-i,P- n) of easily biodegradable soluble microbial by-product substance were higher than others, and little nonbiodegradable fulvic acid-like substance was accumulated. (C) 2016 Elsevier Ltd. All rights reserved.