title: Bioanalytical validation for simultaneous quantification of non-aromatic steroids in follicular fluid from cattle via ESI-LC-MS/MS.
abstract: The family of steroid hormones is quite attractive for the approach of phenotype monitoring in farm animals. Therefore, we developed a new protocol for the quantitative analysis of natural steroids in follicular fluid from dairy cows. The corresponding steroid profile, which consists of progesterone, corticosterone, hydrocortisone, testosterone, and androstenedione covering three distinct steroid classes, was determined by LC/MS. Quantification is achieved by use of steroid standards diluted in steroid-free follicular fluid as calibrators. Thus, the new protocol does not require deuterated standards. In order to correct for conditional performance of the analytical system we have used dexamethasone as an internal standard. The method was validated according to EMA guidelines. Within- and between-day variations were below 20% for most parameters assessed. All steroids assessed had lower limits of quantification in the range of 2.1 to 4.4ng/ml. We have established a simple and sensitive analytical system in order to step towards a broader and cost-efficient phenotyping analysis in follicular fluid from dairy cows.
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Kunze
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Elisa
authors.initials: E
authors.lastname: Wirthgen
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Walz
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Marion
authors.initials: M
authors.lastname: Spitschak
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Brenmoehl
authors.affiliation: Institute of Reproductive Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Vanselow
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Manfred
authors.initials: M
authors.lastname: Schwerin
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Wimmers
authors.affiliation: Institute of Genome Biology, Leibniz Institute for Farm Animal Biology, W.-Stahl-Allee 2, 18196 Dummerstorf, Germany. Electronic address: hoeflich@fbn-dummerstorf.de.
authors.forename: Andreas
authors.initials: A
authors.lastname: Hoeflich
mesh_terms: D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D015571:Follicular Fluid / Q000737:chemistry*; D057230:Limit of Detection; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013256:Steroids / Q000032:analysis*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D013256:Steroids
keywords: Cattle; ESI-LC–MS/MS; Follicular fluid; Phenotyping; Quantification; Steroid hormones; Validation