Journal of Soils and Sediments

, Volume 11, Issue 2, pp 330–335 | Cite as

Boric acid as alternative reference substance for earthworm field tests

  • Petra SteggerEmail author
  • Klaus Peter Ebke
  • Jörg Römbke
Short Original Communication



Boric acid was applied in an earthworm field test according to ISO 11268-3 as a possible alternative for the currently used reference substances that may no longer be available in the near future.

Material and methods

The test site was a pasture with a silt- and clay-dominated soil, a pH of 5.7 and an organic content of 2.8%. In addition to a water-only control, two separate treatments of boric acid were applied: single applications at rates of 187 and 750 kg/ha. In order to investigate the acute impact of this compound, biological sampling was undertaken 4 weeks after application by hand sorting combined with formaldehyde extraction.

Results and discussion

The earthworm community consisted of seven species commonly found at German grassland sites. In the control plots, mean abundance was very high: 984 individuals per square metre before application and 390 individuals per square metre 4 weeks following application of boric acid. In the treated plots, abundance and biomass were reduced by more than 50% compared to the control plots, respectively. In general, juvenile earthworms as well as the epigeic species were affected most strongly.


On the basis of this study, boric acid has the potential to be an alternative reference substance in assessing the acute impact on earthworm species in field tests.


Boric acid Earthworm Field test ISO 11268-3 Reference substance 



The present study was prepared at the Biodiversity and Climate Research Centre (BiK-F), Frankfurt am Main, and financially supported by the research funding programme “LOEWE—Landes-Offensive zur Entwicklung Wissenschaftlich-ökonomischer Exzellenz” of Hesse’s Ministry of Higher Education, Research, and the Arts.


  1. Bauchhenss J (2005) Zeitliche Veränderungen der Regenwurm-Taxozönosen auf Grünland- und Ackerflächen. In: Bayrische Landesanstalt für Landwirtschaft (ed) Schriftenreihe der LfL: 20 Jahres Boden-Dauerbeobachtung in Bayern, 8/2005, pp 41–48Google Scholar
  2. Benini S, Rypniewski WR, Wilson KS, Mangani S, Ciurli S (2004) Molecular details of urease inhibition by boric acid: insights into the catalytic mechanism. J Am Chem Soc 126:3714–3715CrossRefGoogle Scholar
  3. Beylich A, Graefe U (2007) Lumbriciden in der Boden-Dauerbeobachtung: Darstellung von Referenzbereichen, Baselines und Veränderungstendenzen an Beispielen aus Norddeutschland. In: Umweltbundesamt (ed) Bodenbiologische Bewertung von Boden-Dauerbeobachtungsflächen (BDF) anhand von Lumbriciden, UBA-Texte 34/07, pp 33–53Google Scholar
  4. Bouché M (1977) Strategies lombriciennes. Ecol Bull 25:122–132Google Scholar
  5. Boyle TP, Smilie GM, Anderson JC, Beeson DR (1990) A sensitive analysis of nine diversity and seven similarity indices. Res J Water Pollut Control Fed 62:749–762Google Scholar
  6. De Vette HQM, van Asten JG, Hanstveit AO (2000) A study on the adsorption/desorption of boric acid manufacturing grade to soil particulates in four soil types. TNO study no. IMW-99-9047-02. TNO Nutrition and Food Research, DelftGoogle Scholar
  7. EC (Environment Canada) (2004) Biological test method: tests for toxicity of contaminated soil to earthworms (Eisenia andrei, Eisenia fetida, or Lumbricus terrestris). EPS 1/RM/43. EC, OttawaGoogle Scholar
  8. ECB (European Chemicals Bureau) (2000) IUCLID Dataset, Substance ID: 10043-35-3. European Commission, BrüsselGoogle Scholar
  9. Edwards PJ, Coulson JM (1992) Choice of earthworm species for laboratory tests. In: Greig-Smith PW, Becker H, Edwards PJ, Heimbach F (eds) Ecotoxicology of earthworms. Athenaeum, Newcastle upon Tyne, pp 37–43Google Scholar
  10. Engels M, Ratte HT (1992) Randomisierte Ähnlichkeitsanalyse von Lebensgemeinschaften am Beispiel von Mesokosmos-Versuchen in der Ökotoxikologie. Verh. Gesellschaft f. Ökologie, Band 21, Berlin 1991Google Scholar
  11. Graefe U, Beylich A (2003) Critical values of soil acidification for annelid species and the decomposer community. Proc. 5th Int. Coll. Enchytraeidae. Newsl Enchytraeidae 8:51–55Google Scholar
  12. Graff O (1953) Die Regenwürmer Deutschlands. Schriftenreihe Forschungsinstitut Landwirtschaft 7:1–70Google Scholar
  13. ISO (International Organization for Standardization) (1994) Soil quality—determination of pH. ISO 10390. ISO, GenèveGoogle Scholar
  14. ISO (International Organization for Standardization) (1995) Soil quality—determination of organic and total carbon after dry combustion (elementary analysis). ISO 10694. ISO, GenèveGoogle Scholar
  15. ISO (International Organization for Standardization) (1998a) Soil quality—determination of particle size distribution in mineral soil material—method by sieving and sedimentation. ISO 10277. ISO, GenèveGoogle Scholar
  16. ISO (International Organization for Standardization) (1998b) Soil quality—determination of total nitrogen by dry combustion. ISO 13878. ISO, GenèveGoogle Scholar
  17. ISO (International Organization for Standardization) (1999) Soil quality—effects of pollutants on earthworms—part 3: guidance on the determination of effects in field situations. ISO 11268-3. ISO, GenèveGoogle Scholar
  18. Krück S, Joschko M, Schultz-Sternberg R, Kroschewski B, Tessmann J (2006) A classification scheme for earthworm populations (Lumbricidae) in cultivated agricultural soils in Brandenburg, Germany. J Plant Nutr Soil Sci 169:651–660CrossRefGoogle Scholar
  19. Kula H, Kokta C (1992) Side effects of selected pesticides on earthworms under laboratory and field conditions. Soil Biol Biochem 24:1711–1714CrossRefGoogle Scholar
  20. Kula C, Heimbach F, Riepert F, Römbke J (2006) Technical recommendations for the update of the ISO earthworm field test guideline (ISO 11268-3). J Soils Sediment 6:182–186CrossRefGoogle Scholar
  21. Lanno R, Wells J, Conder J, Bradham K, Basta N (2004) The bioavailability of chemicals in soil for earthworms. Ecotoxicol Environ Saf 57:39–47CrossRefGoogle Scholar
  22. Lofs-Holmin A (1986) Occurrence of eleven earthworm species (Lumbricidae) in permanent pastures in relation to soil-pH. Swed J Agric Res 16:161–165Google Scholar
  23. Martinez VG, Prashant KR, Zoran MJ (2006) Asexual reproduction and segmental regeneration, but not morphallaxis, are inhibited by boric acid in Lumbriculus variegatus (Annelida: Clitellata: Lumbriculidae). Hydrobiologia 564:73–86CrossRefGoogle Scholar
  24. Murry FJ (1998) A comparative review of the pharmacokinetics of boric acid in rodents and humans. Biol Trace Elem Res 66:331–341CrossRefGoogle Scholar
  25. Pott R (1995) Die Pflanzengesellschaften Deutschlands. UTB, StuttgartGoogle Scholar
  26. Princz J (ed) (2004) Inter-laboratory validation of environment Canada’s new test methods for measuring soil toxicity using earthworms. Report prepared for the biological methods division. Environment Canada, Ottawa, p 39Google Scholar
  27. Reddy KR, Kayastha AM (2006) Boric acid and boronic acids inhibition of pigeonpea urease. J Enzyme Inhib Med Chem 21:467–470CrossRefGoogle Scholar
  28. Römbke J, Ahtiainen J (2007) The search for the “ideal” soil toxicity test reference substance. IEAM 3:464–466CrossRefGoogle Scholar
  29. Römbke J, Beck L, Förster B, Fründ HC, Horak F, Ruf A, Rosciczweski C, Scheurig M, Woas S (1996) Boden als Lebensraum für Bodenorganismen; bodenbiologische Standortklassifikation, Literaturstudie. Landesanstalt für Umweltschutz Baden-Württemberg, Karlsruhe, pp 59–71Google Scholar
  30. Rundgren S (1975) Vertical distribution of lumbricids in southern Sweden. Oikos 26:299–306CrossRefGoogle Scholar
  31. Shomron N, Ast J (2003) Boric acid reversibly inhibits the second step of pre-mRNA splicing. FEBS Lett 552:219–224CrossRefGoogle Scholar
  32. Sims RW, Gerard BM (1999) Earthworms. In: Kermack DM, Barnes RSK (eds) Synopses of the British fauna (new series), vol 31. Brill/Backhuys, London, p 171Google Scholar
  33. Smith EP (1986) Randomized similarity analysis of multispecies laboratory and field studies. In: El Shaarawi AH, Kwiatkowski RE (eds) Statistical aspects of water quality monitoring. Developments in water science 27. Elsevier, Amsterdam, pp 261–269Google Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Petra Stegger
    • 1
    Email author
  • Klaus Peter Ebke
    • 1
  • Jörg Römbke
    • 2
  1. 1.Institut für Gewässerschutz MESOCOSM GmbH, Forschungszentrum Neu-UlrichsteinHomberg (Ohm)Germany
  2. 2.ECT Oekotoxikologie GmbHFlörsheimGermany

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