title: Size Exclusion High Performance Liquid Chromatography: Re-Discovery of a Rapid and Versatile Method for Clean-Up and Fractionation in Chemical Ecology.
abstract: 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.
authors.affiliation: Institute for Zoology, University of Regensburg, Universitätsstraße 31, D-93053, Regensburg, Germany.
authors.forename: Sergej
authors.initials: S
authors.lastname: Sperling
authors.forename: Stephan
authors.initials: S
authors.lastname: Kühbandner
authors.forename: Katharina C
authors.initials: KC
authors.lastname: Engel
authors.forename: Sandra
authors.initials: S
authors.lastname: Steiger
authors.forename: Johannes
authors.initials: J
authors.lastname: Stökl
authors.forename: Joachim
authors.initials: J
authors.lastname: Ruther
mesh_terms: D000818:Animals; D005591:Chemical Fractionation; D002851:Chromatography, High Pressure Liquid; D001517:Coleoptera / Q000254:growth & development / Q000378:metabolism*; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry; D007605:Juvenile Hormones / Q000032:analysis*; D007814:Larva / Q000254:growth & development / Q000378:metabolism; D014863:Wasps / Q000378:metabolism*
chemical_list: D007605:Juvenile Hormones