title: Use of LC-MS-MS as an alternative to currently available immunoassay methods to quantitate corticosterone in egg yolk and albumen.
abstract: Corticosterone (CORT) is the dominant plasma glucocorticoid in birds. There has been increasing interest in the function of CORT in avian egg yolk and in the potential to use CORT concentrations in eggs to quantify stress and to assess the effect of maternal stress on offspring. The concentration of CORT in egg yolk is most frequently assessed using enzyme or radioimmunoassays, alone or in combination with high-performance liquid chromatography. However, the quantification of CORT is frequently hampered by the presence of high concentrations of other steroid hormones which cross-react with the CORT antibody. As an alternative, we developed a sensitive and specific LC-MS-MS method. The sample-preparation procedure consisted of a protein-lipid precipitation step, followed by defatting and clean-up using a C18 SPE column. Chromatography was performed on an Acquity C18 BEH column (50 mm × 2.1 mm i.d., dp: 1.7 μm, run-time: 6 min), using 0.1% formic acid in both water (A) and acetonitrile (B) as mobile phases. The MS-MS instrument was operated in the positive-electrospray-ionization mode. The method was validated in-house according to European Guidelines (linearity, accuracy and precision, limits of quantification and detection, specificity, stability) and the results fell within the accepted ranges. The method was successfully used for the analysis of CORT in yolk and albumen of eggs collected from eight breeding lesser black-backed gulls at a Flemish coastal colony. CORT concentrations were in the range 42.4-166.3 pg g(-1) in albumen and < LOQ (75 pg g(-1))-762.5 pg g(-1) in yolk.
authors.affiliation: Department of Pharmacology, Toxicology and Biochemistry, Laboratory of Pharmacology and Toxicology, Faculty of Veterinary Medicine, Ghent University, 9820, Merelbeke, Belgium, siegrid.debaere@ugent.be.
authors.forename: Siegrid
authors.initials: S
authors.lastname: De Baere
authors.forename: Tom
authors.initials: T
authors.lastname: Rosendahl Larsen
authors.forename: Mathias
authors.initials: M
authors.lastname: Devreese
authors.forename: Patrick
authors.initials: P
authors.lastname: De Backer
authors.forename: Liesbeth
authors.initials: L
authors.lastname: De Neve
authors.forename: Graham
authors.initials: G
authors.lastname: Fairhurst
authors.forename: Luc
authors.initials: L
authors.lastname: Lens
authors.forename: Siska
authors.initials: S
authors.lastname: Croubels
mesh_terms: D000818:Animals; D046091:Charadriiformes; D011232:Chemical Precipitation; D002645:Chickens; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003345:Corticosterone / Q000032:analysis*; D004529:Egg White / Q000737:chemistry*; D004530:Egg Yolk / Q000737:chemistry*; D007118:Immunoassay; D057230:Limit of Detection; D052616:Solid Phase Extraction / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D003345:Corticosterone