Applied Microbiology and Biotechnology

, Volume 63, Issue 2, pp 217–221 | Cite as

Improved method for determination of ammonia and nitrite oxidation activities in mixed bacterial cultures

  • M. S. Moussa
  • H. J. Lubberding
  • C. M. Hooijmans
  • M. C. M. van Loosdrecht
  • H. J. Gijzen
Short Contribution


A simple and reliable method to measure the activity of ammonia and nitrite oxidisers in mixed bacterial cultures was developed. The developed method differentiates between the ammonia and nitrite oxidisers by consecutive injection of NO2 and NH4+. The main advantage of this method is that it avoids the use of metabolic inhibitors for ammonia or nitrite oxidisers, as used by other methods. Moreover, it allows measuring of the short-term effect of an inhibitor on both the ammonia and nitrite oxidisers in one test under controlled environmental conditions (pH, temperature). The developed method was applied to determine the inhibitory effects of salt (NaCl up to 15 g Cl/l) on an enriched culture of nitrifying bacteria. The results of the method demonstrate its potential to accurately determine the individual activities of nitrite and ammonia oxidisers.


  1. Anthonisen AC, Loher RC, Prakasam TBS, Srinath EG (1976) Inhibition of nitrification by ammonia and nitrous acid. J Water Pollut Control Fed 48:835–852Google Scholar
  2. Antoniou P, Hamilton J, Koopman B, Jain R, Holloway B, Lyberatos G, Svoronos SA (1990) Effect of temperature and pH on the effective maximum specific growth rate of nitrifying bacteria. Water Res 24:97–101Google Scholar
  3. APHA (1995) Standard methods for the examination of water and wastewater, 19th edn. American Public Health Association/American Water Works Association/Water Environment Federation, Washington D.C.Google Scholar
  4. Belser LW, Mays EL (1980) Specific inhibition of nitrite oxidation by chlorate and its use in assessing nitrification in soil and sediments. Appl Environ Microbiol 39:505–510Google Scholar
  5. Ginestet P, Audic JM, Urbain V, Block JC (1998) Estimation of nitrifying bacterial activities by measuring oxygen uptake rate in the presence of the metabolic inhibitor allylthiouria and azide. Appl Environ Microbiol 64:2266–2268PubMedGoogle Scholar
  6. Ginkel CG van, Tramper J, Luyben KChAM, Klapwijk A (1983) Characterisation of Nitrosomonas europaea immobilised in calcium alginate. Enzyme Microb Technol 5:297–303CrossRefGoogle Scholar
  7. Hellinga C, Schellen AAJC, Mulder JW, van Loosdrecht MCM, Heijnen JJ (1998) The SHARON process: an innovative method for nitrogen removal from ammonia-rich wastewater. Water Sci Technol 37:135–142Google Scholar
  8. Hunik JH (1993) Engineering aspects of nitrification with immobilised cells. PhD Thesis. Wageningen Agricultural University, The NetherlandsGoogle Scholar
  9. Hynes RK, Knowles R (1983) Inhibition of chemoautotrophic nitrification by sodium chlorate and sodium chlorite: a re-examination. Appl Environ Microbiol 45:1178–1182Google Scholar
  10. Kong Z, Vanrolleghem P, Willems P, Verstraete W (1996) Simultaneous determination of inhibition kinetics of carbon oxidation and nitrification with a respirometer. Water Sci Technol 30:825–836CrossRefGoogle Scholar
  11. Leenen EJTM, Boogert AA, van Lammeren AAM, Tramper J, Wijffels RH (1997) Dynamic of artificially immobilised Nitrosomonas europaea: effect of biomass decay. Biotechnol Bioeng 55:630–641CrossRefGoogle Scholar
  12. Loosdrecht MCM van, Jetten MSM (1998) Microbiological conversions in nitrogen removal. Water Sci Technol 38:1–7CrossRefGoogle Scholar
  13. Nowak O, Svardal K (1993) Observations on the kinetics of nitrification under inhibiting conditions caused by industrial wastewater compounds. Water Sci Technol 28:115–123Google Scholar
  14. Nowak O, Svardal K, Schweighofer P(1995) The dynamic behaviour of nitrifying activated sludge systems influenced by inhibiting wastewater compounds. Water Sci Technol 31:115–124CrossRefGoogle Scholar
  15. Reichert P, Ruchti J, Simon W (1994) Aquasim 2.0. Swiss Federal Institute For Environmental Science and Technology (EAWAG), Dübendorf, SwitzerlandGoogle Scholar
  16. Smolders GJF, van Loosdrecht MCM, Heijnen JJ (1994) Stoichiometric model of the aerobic metabolism of the biological phosphorus removal process. Biotechnol Bioeng 44:837–848Google Scholar
  17. Spanjers H, Vanrolleghem P, Olsson G, Dold PL (1998) Respirometry in control of the activated sludge process: principles. Int Water Assoc Q, Scientific and Technical Report 7Google Scholar
  18. Sumacz-Gorska J, Gernaey K, Demuynck C, Vanrolleghem P, Verstraete W (1995) Nitrification process control in activated sludge using oxygen uptake measurements. Environ Technol 16:569–577Google Scholar
  19. Sumacz-Gorska J, Gernaey K, Demuynck C, Vanrolleghem P, Verstraete W (1996) Nitrification monitoring in activated sludge by oxygen uptake (OUR) measurements. Water Res 30:1228–1236CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  • M. S. Moussa
    • 1
  • H. J. Lubberding
    • 1
  • C. M. Hooijmans
    • 1
  • M. C. M. van Loosdrecht
    • 2
  • H. J. Gijzen
    • 1
  1. 1.UNESCO-IHE Institute for Water EducationDelftThe Netherlands
  2. 2.Kluyver Institute for BiotechnologyDelft-TUDelftThe Netherlands

Personalised recommendations