title: Combined liquid chromatography-tandem mass spectrometry analysis of progesterone metabolites.
abstract: Progesterone has a number of important functions throughout the human body. While the roles of progesterone are well known, the possible actions and implications of progesterone metabolites in different tissues remain to be determined. There is a growing body of evidence that these metabolites are not inactive, but can have significant biological effects, as anesthetics, anxiolytics and anticonvulsants. Furthermore, they can facilitate synthesis of myelin components in the peripheral nervous system, have effects on human pregnancy and onset of labour, and have a neuroprotective role. For a better understanding of the functions of progesterone metabolites, improved analytical methods are essential. We have developed a combined liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for detection and quantification of progesterone and 16 progesterone metabolites that has femtomolar sensitivity and good reproducibility in a single chromatographic run. MS/MS analyses were performed in positive mode and under constant electrospray ionization conditions. To increase the sensitivity, all of the transitions were recorded using the Scheduled MRM algorithm. This LC-MS/MS method requires small sample volumes and minimal sample preparation, and there is no need for derivatization. Here, we show the application of this method for evaluation of progesterone metabolism in the HES endometrial cell line. In HES cells, the metabolism of progesterone proceeds mainly to (20S)-20-hydroxy-pregn-4-ene-3-one, (20S)-20-hydroxy-5α-pregnane-3-one and (20S)-5α-pregnane-3α,20-diol. The investigation of possible biological effects of these metabolites on the endometrium is currently undergoing.
authors: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors: Maša
authors: M
authors: Sinreih
authors: Institute of Experimental Genetics, Genome Analysis Centre, Helmholtz Zentrum München, München, Germany.
authors: Sven
authors: S
authors: Zukunft
authors: Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia.
authors: Izidor
authors: I
authors: Sosič
authors: Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia.
authors: Jožko
authors: J
authors: Cesar
authors: Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia.
authors: Stanislav
authors: S
authors: Gobec
authors: Institute of Experimental Genetics, Genome Analysis Centre, Helmholtz Zentrum München, München, Germany; Lehrstuhl für Experimentelle Genetik, Technische Universität München, Freising-Weihenstephan, Germany; German Center for Diabetes Research, Neuherberg, Germany.
authors: Jerzy
authors: J
authors: Adamski
authors: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors: Tea
authors: T
authors: Lanišnik Rižner
mesh_terms: D002460:Cell Line; D002853:Chromatography, Liquid; D004717:Endometrium / Q000378:metabolism; D005260:Female; D006801:Humans; D055442:Metabolome; D055432:Metabolomics / Q000379:methods; D011247:Pregnancy; D011374:Progesterone / Q000378:metabolism*; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry
chemical_list: D011374:Progesterone