Composition and Distribution of Extracellular Polymeric Substances in Aerobic Flocs and Granular Sludge

TLDR

The study compared several EPS extraction methods to assess how they influence chemical characterization and used fluorescent stains to visualize EPS in intact samples and cryosections. EPS was quantified in flocculent and aerobic granular sludge from two sequencing batch reactors with identical shear but different settling times, and the study employed multiple extraction methods and fluorescent staining of intact samples and 20‑µm cryosections. Reactor 1 produced flocculent sludge with an SVI of about 120 ml g⁻¹, whereas Reactor 2 formed compact granules with an SVI of.

Abstract

ABSTRACT Extracellular polymeric substances (EPS) were quantified in flocculent and aerobic granular sludge developed in two sequencing batch reactors with the same shear force but different settling times. Several EPS extraction methods were compared to investigate how different methods affect EPS chemical characterization, and fluorescent stains were used to visualize EPS in intact samples and 20-μm cryosections. Reactor 1 (operated with a 10-min settle) enriched predominantly flocculent sludge with a sludge volume index (SVI) of 120 ± 12 ml g −1 , and reactor 2 (2-min settle time) formed compact aerobic granules with an SVI of 50 ± 2 ml g −1 . EPS extraction by using a cation-exchange resin showed that proteins were more dominant than polysaccharides in all samples, and the protein content was 50% more in granular EPS than flocculent EPS. NaOH and heat extraction produced a higher protein and polysaccharide content from cell lysis. In situ EPS staining of granules showed that cells and polysaccharides were localized to the outer edge of granules, whereas the center was comprised mostly of proteins. These observations confirm the chemical extraction data and indicate that granule formation and stability are dependent on a noncellular, protein core. The comparison of EPS methods explains how significant cell lysis and contamination by dead biomass leads to different and opposing conclusions.