Analysis of phase composition in aqueous two-phase systems using a two-column chromatographic method: Application to...

A new chromatographic system for the simultaneous analysis of polyethylene glycol, dextran, sugars, and low-molecular-weight fatty acids was developed. The system is based on a gel exclusion column which allows a first separation between high- and low-molecular-weight compounds, and a cationic exchange column used to further separate the low-molecular-weight compounds. Two applications of the system were demonstrated: (i) after optimizing eluent conditions the gel exclusion column was used to determine the influence of lactic acid, phosphate buffer, and lactic acid bacteria on the ethylene oxide propylene oxide-dextran T40 phase diagram by HPLC; (ii) the ion exchange column was coupled in series with the gel exclusion column and the concentration of polyethylene glycol, dextran, glucose, lactate, acetate, and formate was determined in samples from the fermentative production of lactic acid in a polyethylene glycol 8000-dextran T40 aqueous two-phase system. The fermentation was operated without pH control in a repeated extractive batch mode, where the cell-free top phase was replaced four times, whereas the cell-containing bottom phase was reused repeatedly. The yield was 1.1 mol of lactic acid formed per mole of glucose added and the productivity was 4.7 mM.h(-1). The polymeric composition of the fermentation system was monitored during the five repeated extractive batches, and it showed a progressive depletion in polyethylene glycol and a progressive enrichment in dextran. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 54: 303-311, 1997.