Quantification of Biologically and Chemically Bound Phosphorus Inactivated Sludge from Full-Scale Plants with Biological P-Removal

:phosphorus (P) is present in activated sludge from wastewatertreatment plants in the form of metal salt precipitates, extracellular polymericsubstances, or bound into the biomass, for example, as intracellularpolyphosphate (poly-P). Several methods for a reliable quantification of thedifferent P-fractions have recently been developed, and this study combinesthem to obtain a comprehensive P mass-balance of activated sludge from fourenhanced biological phosphate removal (EBPR) plants. Chemical character-ization by ICP-OES and sequential P fractionation showed that chemicallybound P constituted 38-69% of total P, most likely in the form of Fe, Mg, or Alminerals. Raman microspectroscopy, solution state31P NMR, and31P MASNMR spectroscopy applied before and after anaerobic P-release experiments,were used to quantify poly-P, which constituted 22-54% of total P and wasfound in approximately 25% of all bacterial cells. Raman microspectroscopy incombination withfluorescence in situ hybridization was used to quantify poly-P in known polyphosphate-accumulating organisms(PAO) (Tetrasphaera,CandidatusAccumulibacter, andDechloromonas) and other microorganisms known to possess high level ofpoly-P, such as thefilamentousCa.Microthrix. Interestingly, only 1-13% of total P was stored by unidentified PAO, highlightingthat most PAOs in the full-scale EBPR plants investigated are known