title: Fast separation and quantification of steroid hormones Δ4- and Δ7-dafachronic acid in Caenorhabditis elegans.
abstract: Separation of isomeric molecular species, e.g. double bond position isomers, is a challenging task for liquid chromatography. The two steroid hormones Δ4- and Δ7-dafachronic acid (DA) represent such an isomeric pair. DAs are 3-ketosteroids found in the nematode Caenorhabditis elegans and generated from cholesterol. Δ4- and Δ7-DA have important biological activities and are produced by two different biological pathways in C. elegans. Here we have described a fast separation method for these two isomers using a 1.3 μm core-shell particle in less than 10 min together with a simple MeOH extraction. Using this method we were able to independently quantify Δ4- and Δ7-DA in C. elegans independently from each other and limits of detection of about 5 ng/ml for each isomer were achieved with a good day-to-day reproducibility. As proof-of-principle the method has been applied to the quantification of DAs in worms fed ad libitum or under bacterial deprivation.
authors.affiliation: Research Unit Analytical BioGeoChemistry, Department of Environmental Sciences, Helmholtz Zentrum München, Ingolstädter Landstraße 1, 85764 Neuherberg, Germany. Electronic address: michael.witting@helmholtz-muenchen.de.
authors.forename: Michael
authors.initials: M
authors.lastname: Witting
authors.affiliation: Research Unit Analytical BioGeoChemistry, Department of Environmental Sciences, Helmholtz Zentrum München, Ingolstädter Landstraße 1, 85764 Neuherberg, Germany.
authors.forename: Hans-Christian
authors.initials: HC
authors.lastname: Rudloff
authors.affiliation: Ecole Normale Supérieure de Lyon - CNRS - Université de Lyon Claude Bernard - Molecular Biology of the Cell Laboratory/UMR5239 46, Allee d'Italie, 69364 Lyon cedex 07, France.
authors.forename: Manjunatha
authors.initials: M
authors.lastname: Thondamal
authors.affiliation: Ecole Normale Supérieure de Lyon - CNRS - Université de Lyon Claude Bernard - Molecular Biology of the Cell Laboratory/UMR5239 46, Allee d'Italie, 69364 Lyon cedex 07, France.
authors.forename: Hugo
authors.initials: H
authors.lastname: Aguilaniu
authors.affiliation: Research Unit Analytical BioGeoChemistry, Department of Environmental Sciences, Helmholtz Zentrum München, Ingolstädter Landstraße 1, 85764 Neuherberg, Germany; Chair of Analytical Food Chemistry, Technische Universität München, Alte Akademie 10, D-85354 Freising-Weihenstephan, Germany.
authors.forename: Philippe
authors.initials: P
authors.lastname: Schmitt-Kopplin
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000737:chemistry*; D002782:Cholestenes / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification*; D007536:Isomerism; D057230:Limit of Detection; D016014:Linear Models; D015203:Reproducibility of Results
chemical_list: D002782:Cholestenes; C533799:delta4-dafachronic acid; C533798:delta7-dafachronic acid
keywords: Caenorhabditis elegans; Dafachronic acid; Isomer separation; Sub-2μm core–shell particles