Application of cross-flow ultrafiltration for isolating exopolymeric substances from a marine diatom (Amphora sp.)
Limnol. Oceanogr. Methods 7:419-429 (2009) | DOI: 10.4319/lom.2009.7.419
ABSTRACT: The increasing recognition of the roles that exopolymeric substances (EPS) play in the aquatic environment necessitates obtaining sufficient quantities of purified EPS for exploration of its physical, chemical, and biological properties, as well as for quantitative and structural analysis of its composition. For this purpose, three preconcentration/ purification techniques, i.e., 1) ethanol precipitation, 2) stirred-cell ultrafiltration, and 3) crossflow ultrafiltration, followed by stirred-cell diafiltration, were compared for their effectiveness to quantitatively isolate EPS from laboratory cultures of Amphora sp. The results showed that the classical ethanol precipitation method was not effective in isolating and concentrating EPS from this seawater culture medium. Stirred-cell ultrafiltration appeared best for harvesting EPS from this diatom. However, because of its limitations in terms of time and volume, cross-flow ultrafiltration needed to be first applied, along with some necessary improvements, followed by a three-step cartridge soaking and stirred-cell diafiltration. After cartridge soaking, the yields of the two ultrafiltration methods were comparable. Two different fractions were obtained from EPS of Amphora sp. by anion exchange chromatography and were characterized respectively. Whereas these purified fractions had similar molecular weights of 1000 kDa, their monosaccharide composition was different. In conclusion, cross-flow ultrafiltration followed by stirred-cell diafiltration with additional cartridge washing turned out to be the optimal method for EPS separation, based on time, cost, and yield.