Saccharification is an energy intensive process of particular industrial interest in agribusiness. Considerable improvements to the energy costs of processing can be made by the elimination of multiple high temperature steps during liquefaction. To facilitate technology development for more energy efficient saccharification, the effectiveness of thermoseparating polymer-based aqueous two-phase reactor systems (ATPRS) in the enzymatic hydrolysis of starch was investigated. The partition behavior of pure a-amylase, and a recombinant, thermostable alpha-amylase (MJA1) from the hyperthermophile, Methanococcus jannaschii and amyloglucosidase in PEO-PPO/salt aqueous two-phase systems was evaluated. All of the studied enzymes partitioned unevenly in these systems. Hydrolysis of soluble starch and corn starch into glucose by thermostable alpha-amylase and amyloglucosidase was performed in an ATPRS coupled with temperature-induced phase separation. Use of the ATPRS reduced the hydrolysis time to half of that for single-phase processing, thus reducing energy inputs. The hydrolysis time was 18 hours for 20% soluble starch with an amyloglucosidase concentration of 2.18 unit mL(-1) in the aqueous system, but only 11 hours in aqueous two-phase reactor system. The extent of product inhibition is greatly reduced. These results reveal the potential for polymer-enhanced extractive bioconversion of starch as a practical technology.