UCON-BENZOYL DEXTRAN AQUEOUS 2-PHASE SYSTEMS - PROTEIN-PURIFICATION WITH PHASE COMPONENT RECYCLING

Benzoyl dextran with a degree of substitution of 0.18 was synthesized by reacting dextran T500 with benzoyl chloride. A new type of aqueous two-phase system composed of benzoyl dextran as bottom phase polymer a nd the random copolymer of ethylene oxide and propylene oxide (Ucon 50 -KB-5100) as top phase polymer has been formed. The phase diagram for the system Ucon 50-HB-5100-benzoyl dextran with a degree of substituti on of 0.18 was determined at room temperature. This two-phase system h as been used to purify 3-phosphoglycerate kinase from baker's yeast. T he top-phase polymer (Ucon) can be separated from target enzyme by inc reasing the temperature. The bottom-phase polymer (benzoyl dextran) co uld be recovered by addition of salt. Yeast homogenate was partitioned in a primary Ucon 50-HB-5100-benzoyl dextran aqueous two-phase system . After phase separation the top phase was removed and temperature-ind uced phase separation was used for formation of a water phase and a Uc on-rich phase. The benzoyl dextran-enriched bottom phase from the prim ary system was diluted, and the polymer was separated from water by ad dition of Na2SO4.