Aqueous two-phase extraction using thermo separating polymer: a new system for the separation of endo-polygalacturonase

作者：Pereira, M; Wu, YT; Venancio, A; Teixeira, J^*

来源：Biochemical Engineering Journal, 2003, 15(2): 131-138.

DOI：10.1016/S1369-703X(02)00190-0

• downstream processing
• aqueous two-phase
• separation
• purification
• thermoseparating
• enzymes

Aqueous two-phase partitioning of endo-polygalacturonase (endo-PG) produced by Kluyveromyces marxianus strains was studied, on systems containing the thennoseparating polymer Ucon 50-HB-5100 (a random copolymer of 50% ethylene oxide (EO) and 50% propylene oxide (PO)) as one of the phase-forming compounds. Ucon 50-HB-5100 (Ucon)-polyvinyl alcohol (PVA 10,000), Ucon 50-HB-5100-hydroxypropyl starch (Reppal PES100) and Ucon 50-HB-5100-(NH4)(2)SO4 systems were tested. Ucon recycling was also investigated. Ucon-(NH4)(2)SO4 system proved to be the most efficient system for aqueous two-phase extraction (ATPE) of endo-PG as when compared to total protein partition, the enzyme was strongly partitioned to the salt-rich phase.
Using Ucon-(NH4)(2)SO4 system, a separation scheme consisting of three stages was proposed and tested at laboratory scale. In the first and second stages, operated in series, extractions were performed using the same Ucon-(NH4)(2)SO4 system. Ucon recycling was done in each stage. In the third stage, removal of the Ucon and salt residues present in the enzyme-rich phase of the second extraction stage was done. The proposed scheme allowed for a 10-fold enzyme concentration and a purification factor close to the expected maximum while maintaining more than 95% of the initial enzyme activity.

• 出版日期2003-8