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

Francesca Petriglieri, Jette F. Petersen, Miriam Peces, Marta Nierychlo, Kamilla Hansen, Cecilie E. Baastrand, Ulla Gro Nielsen, Kasper Reitzel, Per Halkjær Nielsen*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

13 Citations (Scopus)
53 Downloads (Pure)


Phosphorus (P) is present in activated sludge from wastewater treatment plants in the form of metal salt precipitates, extracellular polymeric substances, or bound into the biomass, for example, as intracellular polyphosphate (poly-P). Several methods for a reliable quantification of the different P-fractions have recently been developed, and this study combines them to obtain a comprehensive P mass-balance of activated sludge from four enhanced biological phosphate removal (EBPR) plants. Chemical characterization by ICP-OES and sequential P fractionation showed that chemically bound P constituted 38-69% of total P, most likely in the form of Fe, Mg, or Al minerals. Raman microspectroscopy, solution state 31P NMR, and 31P MAS NMR spectroscopy applied before and after anaerobic P-release experiments, were used to quantify poly-P, which constituted 22-54% of total P and was found in approximately 25% of all bacterial cells. Raman microspectroscopy in combination with fluorescence in situ hybridization was used to quantify poly-P in known polyphosphate-accumulating organisms (PAO) (Tetrasphaera, Candidatus Accumulibacter, and Dechloromonas) and other microorganisms known to possess high level of poly-P, such as the filamentous Ca. Microthrix. Interestingly, only 1-13% of total P was stored by unidentified PAO, highlighting that most PAOs in the full-scale EBPR plants investigated are known.

Original languageEnglish
JournalEnvironmental Science and Technology
Issue number8
Pages (from-to)5132-5140
Number of pages9
Publication statusPublished - 2022

Bibliographical note

Funding Information:
We thank Susanne Bielidt for assistance with cell counting and microscopical analysis. The project was funded by the Villum Foundation (Dark Matter, grant 13351) and Innovation Fund Denmark (ReCoverP, grant 4106-00014B).

Publisher Copyright:
© 2021 American Chemical Society. All rights reserved.


  • biological enhanced phosphorus removal (EBPR)
  • NMR spectroscopy
  • P mass-balance
  • polyphosphate-accumulating organisms (PAO)
  • Raman microspectroscopy


Dive into the research topics of 'Quantification of Biologically and Chemically Bound Phosphorus in Activated Sludge from Full-Scale Plants with Biological P-Removal'. Together they form a unique fingerprint.

Cite this