Analytical and Bioanalytical Chemistry

, Volume 379, Issue 3, pp 540–543

Rapid quantification of juvenile hormones and their metabolites in insect haemolymph by liquid chromatography–mass spectrometry (LC-MS)

Short Communication

Abstract

A simple, fast and sensitive method was developed for routine determination of juvenile hormone (JH), JH diols and JH acids in insect haemolymph, by liquid chromatography–mass spectrometry (LC-MS). Sample clean-up involves the precipitation of proteins by methanol/isooctane (1:1, v/v), centrifugation and partial evaporation of the organic solvents. Since JH is bound to a carrier protein in the haemolymph, a binding protein (BP) assay was performed to ensure JH is removed during precipitation. The JH compounds were separated on a C18 column (ReproSil-Pur ODS-3) by gradient elution with water and methanol in less than 22 min and analysed by electrospray mass spectrometry. Due to the high abundance of Na+ in insect haemolymph, [M+Na]+ is primarily formed. The limit of detection and quantification was 6 and 20 pg for JHs, and 8 and 25 pg for JH diols, respectively. To demonstrate the applicability of the method to different insect orders, haemolymph samples from the Mediterranean field cricket (Gryllus bimaculatus), the fall armyworm (Spodoptera frugiperda), the pea aphid (Acyrthosiphon pisum) and an ant species (Myrmicaria eumenoides) were analysed.

Keywords

LC-MS Juvenile hormone titer Acyrthosiphon pisum Gryllus bimaculatus Spodoptera frugiperda Myrmicaria eumenoides 

References

  1. 1.
    Wyatt GR, Davey KG (1996) Adv Insect Physiol 26:1–155Google Scholar
  2. 2.
    Ishaaya I (2001) Biochemical sites of insecticide action and resistance. Springer, Berlin Heidelberg New YorkGoogle Scholar
  3. 3.
    Pratt GE, Tobe SS (1974) Life Sci 14:575–586CrossRefPubMedGoogle Scholar
  4. 4.
    Feyereisen R, Tobe SS (1981) Anal Biochem 111:372–375PubMedGoogle Scholar
  5. 5.
    Strambi C, Strambi A, De Reggi ML, Hirn MH, Delaage MA (1981) Eur J Biochem 118:401–406PubMedGoogle Scholar
  6. 6.
    Goodman WG, Coy DC, Baker FC, Xu L, Toong YC (1990) Insect Biochem 20:357–364CrossRefGoogle Scholar
  7. 7.
    Bergot BJ, Ratcliff MA, Schooley DA (1981) J Chromatogr 204:231–244CrossRefGoogle Scholar
  8. 8.
    Rembold H, Lackner B (1985) J Chromatogr 323:355–361CrossRefGoogle Scholar
  9. 9.
    McCormick JP, Schafer TR (1977) J Org Chem 42:387–390Google Scholar
  10. 10.
    Goodmann WG, Adams B (1984) J Chromatogr 294:447–451CrossRefGoogle Scholar
  11. 11.
    Pawliszyn J (2002) Sampling and sample preparation for field and laboratory. Elsevier, AmsterdamGoogle Scholar
  12. 12.
    Evans PD, Wigglesworth VB (1990) Adv Insect Physiol 22:299–396Google Scholar
  13. 13.
    Glinka AV, Braun RP, Edwards JP, Wyatt GR (1995) Insect Biochem Molec Biol 25:775–781CrossRefGoogle Scholar
  14. 14.
    Osborne JC (1986) Methods Enzymol 128:213–222PubMedGoogle Scholar
  15. 15.
    Penefsky H, Tzagoloff A (1972) Methods Enzymol 22:204–219Google Scholar
  16. 16.
    Menke W, Koenig F (1980) Methods Enzymol 69:446–452Google Scholar
  17. 17.
    Peterson RC, Reich MF, Dunn PE, Law JH, Katzenellenbogen JA (1977) Biochemistry 16:2305–2311PubMedGoogle Scholar
  18. 18.
    Goodman W, Schooley DA, Gilbert LI (1978) Proc Natl Acad Sci USA 75:185–189Google Scholar
  19. 19.
    Prestwich GD, Robles S, Wawrzenczyk C, Buehler A (1987) Insect Biochem 74:551–560CrossRefGoogle Scholar
  20. 20.
    Hammock BD (1985) Comprehensive insect physiology, biochemistry and pharmacology. Pergamon Press, OxfordGoogle Scholar
  21. 21.
    Lanzrein B, Wilhelm R, Riechsteiner R (1993) J Insect Physiol 39:53–63CrossRefGoogle Scholar
  22. 22.
    Nappi AJ, Sugumaran M (1993) Insect Immunity. Kluwer, DordrechtGoogle Scholar
  23. 23.
    Okot-Kotber M, Liavoga A, Yong K-J, Bagorogoza K (2002) J Agric Food Chem 50:2410–2417CrossRefPubMedGoogle Scholar
  24. 24.
    Warrington JC, Saville JC (1999) Biotechnol Bioeng 65:325–333CrossRefPubMedGoogle Scholar
  25. 25.
    Croston G, Abdel-Aal YAI, Gee SJ, Hammock BC (1987) Insect Biochem 17:1017–1021CrossRefGoogle Scholar
  26. 26.
    Schooley DA, Baker FC, Tsai LW, Miller CA, Jamieson GC (1984) Biosynthesis, metabolism and mode of action of invertebrate hormones. Springer-Verlag, Berlin Heidelberg New YorkGoogle Scholar
  27. 27.
    Hazarika LK, Gupta AP (2000) Pest Res J 12:258–260Google Scholar

Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  1. 1.Department of Animal Ecology I, University of BayreuthBayreuthGermany

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