Fractionation of Extracellular Polymeric Substances by Aqueous Three-Phase Partitioning Systems.

Extracellular polymeric substances (EPS) are natural polymers secreted by microorganisms and represent a key chemical for the development of a range of circular economy applications. The production of EPS comes with notable challenges such as downstream processing. In this work, a three-phase partitioning (TPP) system was investigated as a fractionation technique for the separation of polysaccharides and proteins, both present in the EPS culture broth. The effect of the type of phase-forming compounds (alcohol, polymer, or ionic liquid, in combination with salt) and its concentration were evaluated and compared to the results previously obtained with model systems. The recyclability of phase-forming compounds used to form the fractionation platform was assessed by ultrafiltration. The best fractionation of EPS was achieved using a TPP system composed of 23 wt % ethanol and 25% K3C6H5O7 as 82% EPS-PS partitioned to the salt-rich/bottom phase, and 76% EPS-PN was recovered as an interfacial precipitate, which could be readily resolubilized in water. This represented an increase of 1.24 and 2.83-fold in the purity of EPS-PS and EPS-PN, respectively, in relation to the initial feed concentration. Finally, high recovery yields of phase-forming compounds (>99%) and fractionated EPS (>80%) were obtained using ultrafiltration/diafiltration (UF/DF) as the regeneration technique. The substantial fractionation yields, selectivity, and recyclability of the phase-forming compounds confirm the potential of TPP systems in combination with UF/DF as the separation method for real biopolymer mixtures. Key contributions of this study include the demonstration of the applicability of a readily scalable and cost-effective separation technique for the fractionation of EPS from real EPS-containing broths, while also the limitations of prescreening with model systems became clear through the observed deviating trends between model system studies and real broth studies.

High-throughput Soxhlet extraction method applied for analysis of leaf lignocellulose and non-structural substances.

The traditional Soxhlet extraction method is commonly employed to extract soluble components from non-soluble components in a solid matrix, for example, non-structural substances in biomass samples that can be separated from structural lignocellulosic compounds in biomass samples. Conventional laboratory procedures for such extractions typically involve a low sample throughput, with each run being performed individually, resulting in time-consuming and labour-intensive processes, making them impractical for analysing large sample sets. In research fields such as Earth Observation in Forest Ecosystems, extensive fieldwork sampling is required across large study areas, resulting in a substantial number of leaf samples, each with limited mass. In this study, an innovative adaptation of the conventional National Renewable Energy Laboratory (NREL) Soxhlet method is developed to create a high-throughput mini-Soxhlet apparatus that enables the simultaneous extraction of up to nineteen samples, each with a mass of 0.3 g per sample. With this adaptation, we measured the lignocellulose and extractive in 343 leaf samples collected from four temperate forest tree species. This modified approach enhances versatility and can be applied to all solid-liquid extractions and various types of vegetation tissues, such as tree leaves, shrubs, crops, feedstock, and other non-woody samples.•The solid-liquid extraction method has been implemented in a heating block facilitating 19 small flasks to measure multiple samples simultaneously while requiring only a small sample mass.•The apparatus set-up was constructed using an alumina heating block mounted on a standard laboratory heating plate. Boiling flask tubes were placed in the heating block and equipped with condenser caps and filters on glass rods on which the solid samples were placed.•The adjustments made the method suitable for application to diverse vegetation tissues and non-woody sample types. It holds particular appeal for research areas that necessitate a high sample number.