Journal of Chemical Ecology

, Volume 41, Issue 6, pp 574–583 | Cite as

Size Exclusion High Performance Liquid Chromatography: Re-Discovery of a Rapid and Versatile Method for Clean-Up and Fractionation in Chemical Ecology

  • Sergej Sperling
  • Stephan Kühbandner
  • Katharina C. Engel
  • Sandra Steiger
  • Johannes Stökl
  • Joachim RutherEmail author


Solvent extraction of bioactive molecules from glands, tissues, or whole organisms is a common first step in chemoecological studies. Co-extraction of a surplus of high boiling materials such as triacylglycerides (TAGs) and other lipids with higher molecular weight might hamper the identification of volatile or medium-volatile semiochemicals by high resolution chromatographic and spectroscopic techniques. Therefore, effective clean-up procedures are needed to separate potential semiochemicals from the accompanying materials. Size exclusion high performance liquid chromatography (SE-HPLC), a technique often disregarded by chemoecologists, has proved to be a rapid and efficient clean-up method for complex crude extracts. We demonstrated that TAGs can be baseline separated from typical semiochemicals within less than 10 min on a porous gel stationary phase based on highly cross-linked polystyrene/divinylbenzene. We applied the method as a rapid one-step clean-up procedure for the analysis of juvenile hormone III in insect hemolymph by gas chromatography–mass spectrometry. We furthermore introduced some recent application examples on insect pheromones to demonstrate that SE-HPLC is not only an effective method for the purification of crude extracts, but can as well be used as a first fractionation step for the bioassay-guided identification of behavior modifying natural products. SE-HPLC can be well operated with low-boiling solvents such as dichloromethane, and results in fraction volumes of typically less than one ml, which decreases the danger of losing volatile analytes during subsequent concentration steps.


Cuticular hydrocarbons Juvenile hormone Pheromones Sample preparation Triacylglycerides 


Conflict of Interest

The authors declare no conflict of interest.


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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Sergej Sperling
    • 1
  • Stephan Kühbandner
    • 1
  • Katharina C. Engel
    • 2
  • Sandra Steiger
    • 2
  • Johannes Stökl
    • 1
  • Joachim Ruther
    • 1
    Email author
  1. 1.Institute for ZoologyUniversity of RegensburgRegensburgGermany
  2. 2.Institute of Evolutionary Ecology and Conservation GenomicsUniversity of UlmUlmGermany

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