Determination of beta-19-nortestosterone and its metabolite alpha-19-nortestosterone in biological samples at the sub parts per billion level by high-performance liquid chromatography with on-line immunoaffinity sample pretreatment.
ABSTRACT An immunoaffinity precolumn (immuno precolumn) packed with Sepharose-immobilized polyclonal antibodies against the anabolic hormone 17 beta-19-nortestosterone (beta-19-NT) was used for the selective on-line pretreatment of raw extracts of urine, bile and tissue samples by high-performance liquid chromatography. Using UV detection (247 nm), beta-19-NT and its metabolite 17 alpha-19-nortestosterone (alpha-19-NT) can be determined in biological samples with a detection limit of 0.05 microgram/kg. Owing to the high clean-up efficiency of the immuno precolumn and the large sample volumes used, confirmation by gas chromatography-mass spectrometry is possible at this level. In urine samples from a calf treated with 19-nortestosterone 17 beta-laurate, the maximum concentrations of beta-19-NT (1.3 micrograms/l) and alpha-19-NT (3.1 micrograms/l) were found seven days after intramuscular administration. In a bile sample from this calf only alpha-19-NT (55 microgram/l) was detected. In meat samples from three treated calves, the concentration of beta-19-NT varied from 0.1 to 1.6 micrograms/kg and no alpha-19-NT could be detected. In liver samples from these calves, the concentrations of beta-19-NT and alpha-19-NT were less than 0.05-0.1 and 0.5-0.9 micrograms/kg, respectively. In the corresponding kidney samples, the concentrations of beta-19-NT and alpha-19-NT were 0.4-0.5 and 0.5-1.6 micrograms/kg, respectively. The application of the same immuno precolumn to the determination of 17 beta- and 17 alpha-trenbolone, two structurally related steroids, is also demonstrated.
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ABSTRACT: A monoclonal antibody (MAb) raised against sulfamethazine (21C7) was applied in an optical biosensor (Biacore Q) to develop a rapid biosensor immunoassay (BIA) for the detection of several sulfonamides in chicken serum. The performance of this MAb was compared with two polyclonal antibodies (PAbs) raised against sulfamethazine (Qflex sulfamethazine binding protein (SBP) and RIKILT 464b). Using these PAbs, the limits of detection (LODs) in 10 times diluted chicken serum were approximately 30 ng ml−1 and the two BIAs were found to be specific for sulfamethazine. Using MAb 21C7, the LOD for sulfamethazine in 10 times diluted chicken serum was lower (10 ng ml−1), high cross-reactivities were measured for sulfisoxazole (149%), sulfachlorpyridazine (112%), sulfachlorpyrazine (94%), sulfamerazine (87%), sulfadiazine (56%), sulfatroxazole (56%) and sulfathiazole (50%) and low cross-reactivities (11–25%) were measured with six other sulfonamides.Compared with the polyclonal antibodies, the MAb-based BIA resulted in a better sensitivity and was found suitable for the detection of 8 sulfonamides in 10 times diluted chicken serum with LODs between 7 and 20 ng ml−1. The total run time for each cycle was 7 min.Analytica Chimica Acta 01/2003; · 4.39 Impact Factor
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ABSTRACT: 17beta-Nandrolone (17beta-NT) is one of the most recurrent forbidden anabolic steroid used in meat producing animals breeding. Because efficient control must both take into account metabolic patterns and associated kinetics of elimination, the metabolism of 17beta-NT in bovines has already been investigated and is well documented, but only focussing on its main metabolites (i.e. 17alpha-nandrolone, 19-noretiocholanolone and 19-norandrostenedione). The goal of the present study was to enlarge this panel of 17beta-NT metabolites, especially through the urinary estranediols fraction in order to perform a more global steroid profiling upon 17beta-nortestosterone laureate ester administration in calves. A GC-MS/MS method has been developed to monitor and quantify 5 estranediols isomers including 5alpha-estrane-3beta,17beta-diol (abb), 5beta-estrane-3alpha,17beta-diol (bab), 5alpha-estrane-3beta,17alpha-diol (aba), 5alpha-estrane-3alpha,17beta-diol (aab) and 5beta-estrane-3alpha,17alpha-diol (baa). Their urinary elimination kinetics have been established allowing detection of 4 estranediols up to several days after administration. All animals demonstrated homogeneous patterns of elimination both from a qualitative (metabolite profile) and quantitative point of view (elimination kinetics in urine). 5alpha-Estrane-3beta,17alpha-diol (aba) was found as the major metabolite with concentrations up to 100microgL(-1).The Journal of steroid biochemistry and molecular biology 08/2010; 121(3-5):626-32. · 3.98 Impact Factor
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ABSTRACT: Three different group-specific anti-sulfonamide antibodies were compared in inhibition assay formats in an optical biosensor (BIACORE 3000) using CM5 sensor chips coated with three different sulfonamide derivatives. The antibodies used were an anti-sulfamethazine monoclonal antibody (Mab) 21C7, the sulfonamide binding protein (SBP) in the Qflex Kit Sulfonamides and a recently developed mutant antibody (M.3.4). Each of these antibodies showed interactions with all 17 sulfonamides tested and one (Mab 21C7) was sensitive for the N4-acetyl metabolites also. The limits of detection of the different sulfonamides in chicken serum varied between 7 and >1000 ng ml−1 (Mab 21C7), 15 and 340 ng ml−1 (Qflex) and 4 and 82 ng ml−1 (Mutant M.3.4). The mutant M.3.4 based assay was found to be the most sensitive towards most of the sulfonamides whereas the Qflex Kit Sulfonamides detected the five sulfonamides registered for application in poultry in The Netherlands within the narrowest measurement range.Analytica Chimica Acta. 01/2005;