Screening of eltanolone metabolites in dog urine by anion-exchange/reversed-phase liquid chromatography and mass spectrometry.
Strong anion-exchange (SAX) chromatography and reversed-phase liquid chromatography (RPLC) followed by different mass spectrometric techniques for the separation and identification of conjugated and unconjugated 14C-labelled eltanolone (5beta-Pregnan-3alpha-ol-20-one) metabolites in biological fluids are presented. Conjugates of estradiol were used as model compounds for the development of a SAX based group separation of neutral steroids, glucuronides, sulfates and di-conjugated steroids. The usefulness of the technique is demonstrated by the analysis of 14C-labelled eltanolone metabolites in dog urine. The analytical SAX column used prior to RPLC improved the capacity to separate the metabolites from each other and from endogenous components, compared to a single reversed-phase system. Liquid chromatography negative ion electrospray-mass spectrometry (LC-ESI-MS) was used for the molecular mass determination of conjugated eltanolone metabolites. Unconjugated metabolites and hydrolysed conjugates were identified using gas chromatography-mass spectrometry with an electron impact ion source (GC-MS) after trimethylsilyl (TMS) derivatization. An unexpected finding in dog urine was the diglucuronide formation of eltanolone (presumably after enolisation of its carbonyl group).
Tjernberg
A
A
Department of Structural Chemistry, Pharmacia and Upjohn, Stockholm, Sweden.
Edlund
P O
PO
Norén
B
B
D000818:Animals; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004285:Dogs; D005260:Female; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D015394:Molecular Structure; D011280:Pregnanolone / Q000737:chemistry / Q000652:urine*; D013056:Spectrophotometry, Ultraviolet
D011280:Pregnanolone