Confirmatory analysis of 17beta-boldenone, 17alpha-boldenone and androsta-1,4-diene-3,17-dione in bovine urine by liquid chromatography-tandem mass spectrometry.
A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for confirmatory analysis of 17beta-boldenone (17beta-BOL), 17alpha-boldenone (17alpha-BOL) and androsta-1,4-diene-3,17-dione (ADD) in bovine urine was developed. [2H(2)]17beta-Testosterone (17beta-T-d(2)) was used as the internal standard. Sample preparation involved enzymatic hydrolysis and purification on a C(18) solid-phase extraction column. Chromatographic separation of the analytes was obtained using an RP-C(18) HPLC column. LC-MS-MS detection was carried out with an atmospheric pressure chemical ionisation (APCI) source equipped with a heated nebulizer (HN) interface operating in the positive ion mode. For unambiguous hormone confirmation, three analyte precursor-product ion combinations were monitored during multiple-reaction monitoring (MRM) LC-MS-MS analysis. Overall recovery (%), repeatability (relative standard deviations, RSD, %) and within-laboratory reproducibility (RSD, %) ranged from 92.2 to 97.7%, from 6.50 to 2.94% and from 13.50 to 5.04%, respectively, for all analytes. The limit of quantification in bovine urine was 0.20 ng ml(-1) for 17beta-BOL and ADD and 0.50 ng ml(-1) for 17alpha-BOL. The validated method was successfully applied for determination of 17beta-BOL, 17alpha-BOL and ADD in a large number of bovine urine samples collected within the national Official Residue Control Program.
Laboratorio di Medicina Veterinaria, Istituto Superiore di Sanità, V.le Regina Elena, 299, 00161, Rome, Italy. draisci@iss.it
Rosa
R
Draisci
Luca
L
Palleschi
Emanuele
E
Ferretti
Luca
L
Lucentini
Fernanda
F
delli Quadri
D000730:Androstadienes / Q000652:urine*; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013739:Testosterone / Q000031:analogs & derivatives* / Q000652:urine*
D000730:Androstadienes; C006573:1,4-androstadiene-3,17-dione; D013739:Testosterone; C013915:boldenone