Elucidating algal extracellular polymeric substance structures with asymmetrical flow field-flow fractionation and light scattering

Abstract

Extracellular polymeric substances (EPS) from algae are complex, secreted, aquatic heteropolymers (comprised of carbohydrate and proteins), possibly functioning as carbon sinks. EPS has tremendous potential to be utilized as high-value coproducts, e.g. hydrocolloids or biobased polymers, and playing a significant role in the overall aquatic ecology (feeding a healthy microbiome) during cultivation. Unfortunately, the structural elucidation of these polymers is elusive in literature making the design of custom applications difficult. We must characterize these polymers on a chemical, structural, and physical level to understand their biological significance and industrial potential. The first step is to reduce the complexity of EPS with a size-based separation such as asymmetrical flow field-flow fractionation (AF4). When coupled to multi-angle light scattering (MALS), AF4 can provide the separation and characterization needed to determine the molecular weight and size of different populations in the sample. This work evaluates the different size populations present in the EPS of Chlorella vulgaris using AF4-MALS. Fractions were collected and analyzed to probe differences in compositional analyses between the different size populations. The separation investigates aggregate behavior at different ionic strengths to better understand the interactions of these biopolymers in their native, higher salinity, environments. The EPS of C. vulgaris has demonstrated diverse molecular weight populations ranging from 4x104 – 3x108 Daltons. We observed a reduction of fractogram features at high ionic strength indicating polymer aggregation. This work aims to be the first step in complete structural determination of EPS while probing fundamental separation observations on polymer behavior at different salt concentrations.