Boldenone [HRP]

Bioassays are valuable tools for combating the illegal use of steroids in cattle fattening. Previously we described the construction and properties of a rapid and robust yeast androgen bioassay stably expressing the human androgen receptor (hAR) and yeast enhanced green fluorescent protein (yEGFP), the latter in response to androgens. In the present study this yeast androgen bioassay was validated as a qualitative screening method for the determination of androgenic activity in calf urine and animal feed. This validation was performed according to EC Decision 2002/657. 20 blank samples were spiked with testosterone, 17 alpha-methyltestosterone, 19-nortestosterone, 17 beta-trenbolone, 17 beta-boldenone or 17 beta-methylboldenone at 2 or 15 ng mL(-1) in urine and 50 or 100 ng g(-1) in feed. All blank and spiked samples fulfilled the CC alpha and CC beta criterions, meaning that all 20 blank samples gave signals below the determined decision limits CCa and were thus classified as compliant (alpha = 1%). For each component, at least 19 out of the 20 spiked samples gave a signal above the CC alpha and were thus classified as suspect (beta = 5%). The method was specific, and high amounts of dexamethasone did not interfere with the outcome of the test. Although high levels of 17 alpha-ethynylestradiol can significantly inhibit the response obtained with low amounts of androgens, that situation is not relevant in veterinary practice. When stored at their specific conditions, the androgens in feed were stable for at least 91 days. Real urine samples from a national control program were screened and a representative part of the compliant and suspect samples were confirmed by gas chromatography-tandem mass spectrometry. (C) 2008 Elsevier B.V All rights reserved.

A multiresidue analytical method for the determination of the most common and biologically active natural and synthetic steroids (four estrogens: estriol, 17 beta -estradiol, 17 alpha -ethynylestradiol, estrone; one progestagen: progesterone and six androgens: trenbolone, boldenone, nandrolone, testosterone, 17 alpha -methyltestosterone, stanozolol) in environmental waters was developed. The analytes were isolated from water samples by solid phase extraction (SPE) utilizing a graphitized carbon black adsorbent (Carbograph-1). The final samples were analyzed by reversed-phase high performance liquid chromatography with tandem mass spectrometry using atmospheric pressure chemical ionization (LC-APCI-MS-MS). Ionization was performed in a heated nebulizer (HN) interface operating in the positive ion mode. The protonated ions [M+H](+) and the dehydrated ions [M+H-H2O](+) (for estriol, 17 alpha -estradiol, 17 beta -estradiol, and 17 alpha -ethynylestradiol) were used as precursor ion for collision-induced dissociation (CID), and two diagnostic product ions for each analyte were identified for the unambiguous steroid confirmation by multiple reaction monitoring (MRM) mode. Method performance, for this analytical procedure, was validated by analyzing groundwater and river water samples fortified at level of 20 ng/L. The average recovery for each analyte exceeded 82%. Good method precision was demonstrated with percent relative standard deviation of less than 7.2% for all analytes.