Oral administration of boldenone and boldione to veal calves: disposition and elimination rate of boldenone metabolites

INTRODUCTION Boldenone (1,4-androstadiene-3–one, 17-ol) (BOL) and boldione (1, 4-androstadiene-3,17-dione) (ADD) are chemical derivatives of testosterone, known as strong anabolics and low to moderate androgenic agents. ADD, the dione form of BOL, is a direct precursor of that anabolic steroid, and it is activated by the same widely distributed 17beta-hydroxysteroid dehydrogenase enzyme that converts androstenedione to testosterone . ADD is one of the most orally active pro-hormones, and displays a level of oral bioavailability far superior to any other compound. To optimize anabolic surveillance strategies, ADD and BOL were administered to veal calves, at a similar dosage and the same ratio present in ‘ready to use’ cocktails; their disposition in plasma and their elimination rate in urine were followed for 24 h. Analysis Urinary concentrations of b-BOL, a-BOL and ADD were determined (before and after deconjugation with Helix pomatia extracts) using a validated liquid chromatography-tandem mass spectrometry (LC-MS-MS) method. To measure plasma concentrations, the same method with slight modifications was adopted. ADD was not detectable in any of the urine or plasma samples at any time.For both 17 a- and 17b-BOL the highest concentrations were detected in the first plasma samples (15 min after administration), their concentrations then decreased at similar rates and both were undetectable within 4 h after administration. In urine samples collected within 4 h after administration, concentrations of a-BOL were considerably higher than those of b-BOL, and 36 h after administration a-BOL was the only detectable metabolite. b-BOL was detectable over a period of 24 h only. The absence of ADD confirmed its double identity, i.e. not only as a metabolite but also a precursor of b-BOL, as already shown in vitro after incubation of ADD or b-BOL with calf liver microsomes . The in vivo results confirm that both ADD and b-BOL were promptly absorbed when administered to veal calves before feeding, and that their disappearance from plasma was even more rapid. About 3 h after administration a significant quantity of a-BOL was recovered in urine, while b-BOL attained concentrations only slightly higher than the EU action limit. Surprisingly, after 9 h a-BOL concentrations were already less than 2% of those detected at the previous sampling time and b-BOL concentrations were closed to the limit of quantitation. One day after treatment only a-BOL gave a feeble indication of the ‘illegal’ treatment. These findings indicate that sampling time is crucial when urine samples are collected from farmed calves for surveillance purposes.