Decoupling EPS-bound and intracellular phosphorus in waste activated sludge: A low-disturbance strategy for controlled recovery

Abstract

ABSTRACT Phosphorus (P) in waste activated sludge (WAS) exists in both intracellular stores and fractions associated with extracellular polymeric substances (EPS). These EPS-bound phosphorus species are often stabilized through metal – phosphate – EPS linkages, which complicate their selective recovery. In this study, five single extraction methods (acidification, alkalization, heating, ultrasound, and EDTA chelation) and one combined approach (ultrasound-EDTA) were comparatively evaluated to elucidate their mechanisms of EPS disruption and P mobilization. Phosphorus speciation behavior under single treatments was characterized using 31P nuclear magnetic resonance with phytic acid and sodium hexametaphosphate as model compounds, revealing that strong alkaline extraction caused extensive Poly-P depolymerization and OP hydrolysis, whereas EDTA and low-intensity ultrasound preserved P speciation but showed limited extraction efficiency. The ultrasound-EDTA combination achieved a synergistic effect, yielding 62.63 mg·g−1 TSS of EPS and 9.6 mg·g−1 TSS of phosphorus with only 13.2% cell apoptosis, comparable to alkaline extraction yet with minimal biological disruption. Confocal microscopic analysis confirmed that the released phosphorus predominantly originated from the EPS matrix, indicating that the hybrid treatment effectively targeted EPS-P while preventing intracellular release. These findings establish a non-destructive extraction pathway based on the coupling of physical cavitation and selective chelation, providing mechanistic insight into EPS-mediated P binding and offering a new strategy for controlled, high-efficiency phosphorus recovery from sludge. GRAPHICAL ABSTRACTA schematic diagram of sludge extracellular polymeric substances undergoing heat, acid, alkaline, ultrasound, and chelation extraction steps.The figure shows a schematic diagram illustrating how different extraction methods act on sludge extracellular polymeric substances. On the left, a large irregular blob labeled sludge extracellular polymeric substances surrounds a central rounded cluster containing smaller particles and short chain like structures. An arrow points to a vertical sequence of four extraction methods represented by icons and labels: a flame above a flask for heat treatment, a green filled flask for alkaline treatment, a purple filled flask for acid treatment, and an ultrasound probe for ultrasound. Below, text lists ethylenediaminetetraacetic acid chelation and ultrasound plus ethylenediaminetetraacetic acid chelation with a chemical structure of ethylenediaminetetraacetic acid. A long arrow leads to a second, similar sludge extracellular polymeric substances blob on the right, now surrounded by more separated spheres and fragments that represent released components. Around this blob, separate callout boxes describe each method. The heat treatment box includes a thermometer icon, a flame, and scattered spheres and chains with text stating that thermal unfolding weakens hydrogen bonds and partly loosens the extracellular polymeric substances matrix. The acid treatment box shows a purple flask and the letter H plus, with text that proton attack dissolves metal phosphate bonds. The alkaline treatment box shows a green flask and the letters O H minus indicating that this treatment induces extracellular polymeric substance disintegration and polyphosphate hydrolysis, leading to intracellular phosphorus release and cell lysis. Another callout near an ultrasound probe explains that cavitation shear detaches extracellular polymeric substance layers and exposes bound phosphorus while leaving metal phosphate complexes. A further annotation beside ultrasound plus ethylenediaminetetraacetic acid notes that cavitation enhances ethylenediaminetetraacetic acid penetration and metal chelation, producing high extracellular polymeric substance bound phosphorus release with limited polyphosphate hydrolysis. The ethylenediaminetetraacetic acid chelation callout depicts metal chelation as ethylenediaminetetraacetic acid binding to metal ions and states that chelation of iron three plus and aluminum three plus breaks metal bridges. A legend at the bottom right defines three symbols: phosphorus in the extracellular polymeric substances as small internal particles, extracellular polymeric substance as the large blob, and metal ion as solid circles.