Biofilms play important roles in water technologies such as membrane treatments and activated sludge. The extracellular polymeric substances (EPS) are key components of biofilms. However, the precise nature of these substances and how they influence biofilm formation and behavior remain critical knowledge gaps. EPS are produced by many different microorganisms and span multiple biopolymer classes, which each require distinct strategies for characterization. The biopolymers additionally associate with each other to form insoluble complexes. Here, we explore recent progress toward resolving the structures and functions of EPS, where a shift towards direct functional assessments and advanced characterization techniques is necessary. This will enable integration with better microbial community and omics analyses to understand EPS biosynthesis pathways and create further opportunities for EPS control and valorization.

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Over the past decade, a significant amount of research work on extracellular polymeric substances has been done on the “alginate-like exopolysaccharides” (ALE, also called “alginate-like exopolymers”). The term was used based on the FAO (Food and Agriculture Organization) biopolymer identification test. Although various chemical analyses have been conducted to characterize extracted ALE, it remained unclear whether ALE contains the two sugar monomers of alginates. Aiming to obtain a direct answer to the question: are there alginates in the ALE extracted from sludge, activated sludge was collected from two wastewater treatment plants in two different countries, where the ALE was previously extracted, characterized, and reported in the literature. The extracellular polymers were extracted from these sludges and fractionated according to the standard protocol. The sugar monomer composition of each fraction was analyzed, with special attention to the presence of mannuronic acid (M) and guluronic acid (G), which compose alginate biopolymers. None of these monomers were found in the extracted EPS, indicating there are no alginates resembling polymers extracted from the sludges. The possibility of the presence of other glycan components, such as lipopolysaccharides in EPS, was investigated and confirmed.@en

Kaumera are extracellular polymeric substances (EPS) extracted from excess aerobic granular sludge from Nereda® wastewater treatment plants. Kaumera exhibits significant market potential across diverse applications, fostering rapid research and business development. Furthermore, it will begin to be extracted from numerous installations worldwide. This calls for standard methods as analogue to (waste)water and sludge characterization. Due to lack of standardization, stakeholders are currently using different extraction and characterization protocols, impeding the development of a more uniform product and comparison of results across research studies. To address this, this report compiles the standard protocol for Kaumera extraction in the laboratory and for on-site and lab characterization to be used by researchers, the public Dutch water authorities, and the private industry. The procedures detailed in this document are in accordance with EPS research conducted at TU Delft and methodologies employed in Kaumera production facilities. This report aids in monitoring Kaumera characteristics worldwide and for optimizing the extraction process (including up and downstream processing). This will help maximize repeatability, interoperability, and quality and therefore accelerate business and research development, paving the way to develop a product that meets the needs of the endusers. Through the widespread adoption of this manual, our aim is to foster greater coordination and collaboration among stakeholders, thereby expediting the realization of Kaumera's full potential. @en