Biology and Fertility of Soils

, Volume 48, Issue 6, pp 643–650 | Cite as

Evaluation of ecological doses of the nitrification inhibitors 3,4-dimethylpyrazole phosphate (DMPP) and 4-chloromethylpyrazole (ClMP) in comparison to dicyandiamide (DCD) in their effects on dehydrogenase and dimethyl sulfoxide reductase activity in soils

  • Ferisman TindaonEmail author
  • Gero Benckiser
  • Johannes C. G. Ottow
Original Paper


Risk assessment of the nitrification inhibitors (NIs) 3,4-dimethylpyrazole phosphate (DMPP), 4-chloromethylpyrazole (ClMP), and dicyandiamide (DCD) on nontarget microbial activity in soils was determined by measuring dehydrogenase and dimethyl sulfoxide reductase activity (DHA, DRA, respectively) in three differently textured soils under laboratory conditions. Dehydrogenase activity was measured with standard procedure recommended to evaluate side effects of environmental chemicals on general microbial activity in soils. The kinetic of inhibition were obtained by dose–response relationships and used to calculate the no observable effect levels (NOEL values) and the effective doses at 10% and 50% inhibition (ED10 and ED50), respectively. Negative effects on DHA and DRA, respectively, were observed only at rates approximately 40–100 times higher than the concentrations recommended in the field. Both DHA and DRA were affected more in the sandy than in the silty or clayey soil. Consequently, NOEL, ED10, and ED50 values were considerably higher in the clayey than in the silty or sandy soil. The heterocyclic N compounds DMPP and ClMP, respectively, were more effective in inhibiting DHA and DRA than DCD. At application rates used in the field as well as at concentration up to 25 to 90 times higher, the NIs concerned failed to affect general soil microbial activity in soils. Among the three NIs tested, the not marketed ClMP exhibited the strongest negative effects on soil microbial activity. At recommended application rates, the NIs tested should be considered as enviromentally safe.


Nitrification inhibitors Dehydrogenase activity Dimethyl sulfoxide reductase activity No observable effect level (NOEL) ED10 and ED50 



We highly regret that our supervisor and teacher Prof Johannes CG Ottow, who significantly contributed to this paper, unreckoningly died on August 20, 2011. Ferisman Tindaon gratefully acknowledges Prof Dr. JCG Ottow’s cooperativeness and a Ph.D. scholarship provided by Deutscher Akademischer Austauschsdienst (DAAD), Bonn, Germany. This work was supported by the German Ministry for Education and Research (BMBF; Grant Nr. 0339812).


  1. Alef K, Kleiner D (1989) Rapid and sensitive determination of microbial activity in soils and in soil aggregates by dimethylsulfoxide reduction. Biol Fertil Soils 8:349–355CrossRefGoogle Scholar
  2. Alef K, Nanniperi P (1995) Methods in applied soil microbiology and biochemistry. Academic Press, LondonGoogle Scholar
  3. Ali R, Iqbal J, Tahir GR, Mahmood T (2008) Effect of 3,5-dimethylpyrazole and nitrapyrin on nitrification under high soil temperature. Pak J Bot 40:1053–1062Google Scholar
  4. Azam F, Benckiser G, Mueller C, Ottow JCG (2001) Release, movement and recovery of 3,4- dimethylpyrazole phosphate (DMPP), ammonium and nitrate from stabilized fertilizer granules in a silty clay soil under laboratory conditions. Biol Fertil Soils 34:118–125CrossRefGoogle Scholar
  5. Babich H, Bewley RJF, Stotzky G (1983) Application of the ecological dose concept to the impact of heavy metals on some microbe-mediated ecological processes in soil. Arch Environ Contam Toxicol 12:421–426Google Scholar
  6. Bannert A, Mueller-Niggemann C, Kleineidam K, Wissing L, Cao Z, Schwark L, Schloter M (2011) Comparison of lipid biomarker and gene abundance characterizing the archaeal ammonia-oxidizing community in flooded soils. Biol Fertil Soils 47:839–843CrossRefGoogle Scholar
  7. Barth G, von Tucher S, Schmidhalter U (2001) Influence of soil parameter on the effect of 3,4 dimethylpyrazole phosphate (DMPP) as nitrification inhibitor. Biol Fertil Soils 34:98–102CrossRefGoogle Scholar
  8. Barth G, von Tucher S, Schmidhalter U (2008) Effectiveness of 3,4-dimethylpyrazole phosphate as nitrification inhibitor in soil as influenced by inhibitor concentration, application form, and soil matrix potential. Pedosphere 18:378–385CrossRefGoogle Scholar
  9. Benckiser G, Bamforth SS (2010) The role of pathogens, signal recalcitrance, and organisms shifting in ecosystem recuperation. Agron Sustain Dev. doi: 10.1051/agro/2010024
  10. Casida LE Jr (1977) Microbial metabolic activity in soil as measured by dehydrogenase determination. Appl Environ Microbiol 34:630–636PubMedGoogle Scholar
  11. Casida LE Jr, Klein DA, Santoro T (1964) Soil dehydrogenase activity. Soil Sci 98:371–376CrossRefGoogle Scholar
  12. Di HJ, Cameron KC, Sherlock RR (2007) Comparison of the effectiveness of a nitrification inhibitor, dicyandiamide (DCD), in reducing nitrous oxide emissions in four different soils under different climatic and management conditions. Soil Use Manag 23:1–9CrossRefGoogle Scholar
  13. Di HJ, Cameron KC, Shen JP, Winefield CS, O'Callaghan M, Bowatte S, He JZ (2010) Ammonia-oxidizing bacteria and archaea grow under contrasting soil nitrogen conditions. FEMS Microbiol Ecol 72:386–394PubMedCrossRefGoogle Scholar
  14. Fettweis U, Mittelstaedt W, Schimansky C, Fuehr F (2001) Lysimeters experiments on the translocation of the 14C-labelled nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) in a gleyic cambisol. Biol Fertil Soils 34:126–130CrossRefGoogle Scholar
  15. Friedel JK, Mölter K, Fischer WR (1994) Comparison and improvement of methods for determining the dehydrogenase activity of soils with triphenyltetrazolium chloride and iodonitrotetrazolium chloride. Biol Fertil Soils 18:291–296CrossRefGoogle Scholar
  16. Griebler C (1997) Dimethylsulfoxide (DMSO) reduction: an approach to determine microbial activity in freshwater sediment. J Microbiol Meth 29:31–40CrossRefGoogle Scholar
  17. Griebler C, Slezak D (2001) Microbial activity in aquatic environments measured by dimethyl sulfoxide reduction and intercomparison with commonly used methods. Appl Environ Microbiol 67:100–109PubMedCrossRefGoogle Scholar
  18. Haanstra L, Doelman P (1991) An ecological dose-response model approach to short- and long term effects of heavy metals on arylsulphate activity in soil. Biol Fertil Soils 11:18–23CrossRefGoogle Scholar
  19. Haanstra L, Doelman P, Oude Voshaar JH (1985) The use of sigmoidal dose response curves in soil ecological research. Plant Soil 84:293–297CrossRefGoogle Scholar
  20. Hinojosa MB, Carreira JA, García-Ruíz R, Dick RP (2004) Soil moisture pre-treatment effects on enzyme activities as indicators of heavy metal-contaminated and reclaimed soils. Soil Biol Biochem 36:1559–1568CrossRefGoogle Scholar
  21. Irigoyen I, Muro J, Azpilikueta M, Aparicio-Tejo A, Lamsfus C (2003) Ammonium oxidation kinetics in the presence of the nitrification inhibitors DCD and DMPP at various temperatures. Aust J Soil Res 41:1177–1183CrossRefGoogle Scholar
  22. Kostov O, Van Cleemput O (2001) Nitrogen transformation in copper-contaminated soils and effects of line and compost application on soil resiliency. Biol Fertil Soils 33:10–16CrossRefGoogle Scholar
  23. Li H, Xi L, Chen Y, Tian G, Ni W (2008) Effect of nitrification inhibitor DMPP on nitrogen leaching, nitrifying organisms and enzyme activities in a rice-oil seed rape cropping system. J Environ Sci 20:149–155CrossRefGoogle Scholar
  24. Linzmeier W, Gutser R, Schmidhalter U (2001) Nitrous oxide emission from soil and from a nitrogen 15-labelled fertilizer with the new nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP). Biol Fertil Soils 34:103–108CrossRefGoogle Scholar
  25. López-Gutiérrez JC, Toro M, López-Hernández D (2004) Seasonality of organic phosphorus mineralization in the of the native savanna grass, Trachypogon plumosus. Soil Biol Biochem 36:1675–1684CrossRefGoogle Scholar
  26. Mahmood T, Ali R, Latif Z, Ishaque W (2011) Dicyandiamide increases the fertilizer N loss from an alkaline calcareous soil treated with 15N-labelled urea under warm climate and under different crops. Biol Fertil Soils 47:619–631CrossRefGoogle Scholar
  27. Makoi JHJR, Ndakidemi PA (2008) Selected soil enzymes: examples of their potential roles in the ecosystem. Afr J Biotechnol 7:181–191Google Scholar
  28. Mc Carty GW (1999) Modes of action of nitrification inhibitors. Biol Fertil Soils 29:1–9CrossRefGoogle Scholar
  29. McCarty GW, Bremner JM (1989) Inhibition of nitrification in soil by heterocylic nitrogen compounds. Biol Fertil Soils 8:204–211Google Scholar
  30. McCarty GW, Bremner JM (1990) Evaluation of 3-methylpyrazole-1-carboxamide as a soil nitrification inhibitor. Biol Fertil Soils 9:252–256CrossRefGoogle Scholar
  31. Mueller C, Stevens RJ, Laughlin RJ, Azam F, Ottow JCG (2002) The nitrification inhibitor DMPP had no effect on denitrifying enzyme activity. Soil Biol Biochem 34:1825–1827CrossRefGoogle Scholar
  32. Obbard JP (2001) Measurement of dehydrogenase activity using 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride (INT) in the presence of copper. Biol Fertil Soils 33:328–330CrossRefGoogle Scholar
  33. Ottow JCG (2011) Mikrobiologie von Boeden: Biodiversitaet, Oekophysiologie. Metagenomik. Springer, Verlag Berlin-HeidelbergGoogle Scholar
  34. Pasda G, Haehndel R, Zerulla W (2001) Effect of fertilizers with the new nitrification inhibitor DMPP (3,4-dimethylpyrazole phosphate) on yield and quality of agricultural and horticultural crops. Biol Fertil Soils 34:85–97CrossRefGoogle Scholar
  35. Prasad R, Power JF (1995) Nitrification inhibitors for agriculture, health and environment. Adv Agron 54:233–281CrossRefGoogle Scholar
  36. Rajbanshi SS, Benckiser G, Ottow JCG (1992) Effects of concentration, incubation temperature, and repeated applications on degradation kinetics of dicyandiamide (DCD) in model experiments with a silt soil. Biol Fertil Soils 13:61–64CrossRefGoogle Scholar
  37. Rao MA, Sannino F, Nocerino G, Puglisi E, Gianfreda L (2003) Effect of air-drying treatment on enzymatic activities of soils affected by anthropogenic activities. Biol Fertil Soils 38:327–332CrossRefGoogle Scholar
  38. Richter O, Diekkrueger B, Noertersheuser P (1996) Environmental fate and modeling of pesticides from laboratory to field scale. VCH Weinheim, New York, pp 89–93CrossRefGoogle Scholar
  39. Schlichting E, Blume HP, Stahr K (1995) Bodenkundliches Praktikum. Blackwell, BerlinGoogle Scholar
  40. Stephenson GL, Koper N, Atkinson GF, Solomon KR, Scroggins RP (2000) Use of non linear regression techniques for describing concentration response-relationships of plant species exposed to contaminated site soils. Environ Toxicol Chem 19:2968–2981CrossRefGoogle Scholar
  41. USEPA (1984) Technical guidance manual for performing waste load allocations. Book II: stream and rivers. Chapter 3: toxic substances. EPA 440/4-84-022. Office of Water, WashingtonGoogle Scholar
  42. Von Mersi W, Schinner F (1991) An improved and accurate method for determining the dehydrogenase activity of soils with iodonitrotetrazolium chloride. Biol Fertil Soils 11:216–220CrossRefGoogle Scholar
  43. Weiske A, Benckiser G, Ottow JCG (2001a) Effect of the new nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) in comparison to dicyandiamide (DCD) on nitrous oxide (N2O) emission and methane (CH4) oxidation during 3 years repeated application in field experiments. Nutr Cycl Agroecosys 60:57–64CrossRefGoogle Scholar
  44. Weiske A, Benckiser G, Herbert T, Ottow JCG (2001b) Influence of nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) in comparison to dicyandiamide (DCD) on nitrous oxide emission and methane oxidation during 3 years repeated application in field experiments. Biol Fertil Soils 34:109–117CrossRefGoogle Scholar
  45. Wingfield GI, Davies HA, Greaves MP (1977) The effect of soil treatment on the response of the soil microflora to the herbiside dalapon. J Appl Bacteriol 43:39–46CrossRefGoogle Scholar
  46. Zerulla W, Barth T, Dressel J, Erhardt K, Horchler von Locquenqhien K, Pasda G, Raedle M, Wissemeier AH (2001) DMPP—a new nitrification inhibitor for agriculture and horticulture: an introduction. Biol Fertil Soils 34:79–84CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Ferisman Tindaon
    • 1
    • 2
    Email author
  • Gero Benckiser
    • 1
  • Johannes C. G. Ottow
    • 1
  1. 1.Institute for Applied MicrobiologyJustus Liebig UniversityGiessenGermany
  2. 2.Agroecotechnology Department, Faculty of AgricultureNommensen University MedanMedanIndonesia

Personalised recommendations