Extraction and Purification of Exopolysaccharides from Geobacter Biofilms

Exopolysaccharides secreted by Geobacter play a pivotal role in mediating extracellular electron transfer (EET), biofilm formation, and environmental adaptability. However, existing methods for extracting and purifying Geobacter exopolysaccharides often suffer from low yield, structural damage, or contamination by intracellular components, limiting in-depth research on the structure and function of exopolysaccharides. This paper aimed to optimize the extraction and purification protocols for Geobacter exopolysaccharides. Three crude extraction methods (EDTA, high-speed centrifugation, and heating) for exopolysaccharides were evaluated, and the EDTA method was selected as the optimal crude extraction strategy, balancing exopolysaccharides yield (22.3 μg/mL) and cell viability (90.1%), outperforming high-speed centrifugation (lower yield) and heating (severe cell lysis). Purification was optimized using a two-step process: for deproteinization, the Sevag method was optimized to four cycles, removing 75% of proteins with minimal exopolysaccharides loss. Ethanol precipitation was optimized to 75–90% concentration and 24 h incubation, yielding 19.6–20.9 μg/mL of purified exopolysaccharides while eliminating soluble impurities. This optimized protocol ensures high-quality exopolysaccharides isolation with minimal cell lysis and reduced risk of structural disruption, providing a reliable foundation for investigating the roles of Geobacter exopolysaccharides in EET and environmental applications.