Alpha-amylase production in aqueous two-phase systems by Bacillus subtilis

Zoi Konsoula, Maria Liakopoulou-Kyriakides

Research output: Contribution to journalMeeting Abstractpeer-review


The production of a-amylase by Bacillus subtilis was studied in polyethylene glycol (PEG 10 000)/dextran (505 000) aqueous two-phase systems at various concentrations. An increase in the PEG concentration from 7 to 25% (w/w) in the aqueous two-phase system resulted in an increase in the phase volume ratio with a concomitant decrease in the partition coefficient (K) and recovery of a-amylase in the top phase. However, varying dextran concentrations from 2.5 to 10% (w/w) decreased both the phase volume ratio and the partition coefficient of a-amylase. At dextran concentrations lower than 2.5% (w/w) the phases could not separate. The purification ratio was found to increase with increasing PEG concentration up to 9% (w/w), while it was slightly decreased at higher concentrations. The PEG 9% (w/w) and dextran 2.5% (w/w) system was found to be optimum for cultivation of Bacillus subtilis, where more than 95% of the produced a-amylase was selectively partitioned to the upper phase giving a purification factor of 2.3. In this system the a-amylase activity in the top phase, which reached 93 U/ml after 48 h of cultivation, was 1.2 times higher in comparison to the homogeneous medium. It was observed that Bacillus subtilis secreted 84% of the total a-amylase produced within 24 h, while the respective time exceeded 33 h in the homogeneous medium. The bacterial cells were microscopically observed to partition totally to the bottom phase in the system used. The dextran-rich bottom’s phase affinity for cells was exploited in the development of a semicontinuous cell recycle system, in which the bottom phase was reused and the cell-free top phase was withdrawn. In the first two uses a-amylase activity in the top phase exceeded 90 U/ml, while in the third use it was reduced to 84 U/ml. Concluding the use of the PEG 9% (w/w)/dextran 2.5% (w/w) aqueous two-phase system enabled the recirculation of the bacterial cells and the reduction of the required time of cultivation, thus contributing significantly to the reduction of the cost of the fermentation.
Original languageEnglish
Article numberL1-024P
Pages (from-to)497-497
Number of pages1
JournalThe FEBS Journal
Issue numbers1
Publication statusPublished - 2005


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