Development and validation of a liquid chromatography–tandem mass spectrometry method for the separation of conjugated and unconjugated 17α- and 17β-boldenone in urine sample

Gasparini, Mara; Assini, Walter; Bozzoni, Eros; Tognoli, Nadia; Dusi, Guglielmo

doi:10.1016/j.aca.2006.08.059

Cited by 8 publications

(4 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The method used by IZSLER was a fully validated method according to the Commission Decision 2002/657/EC with SPE extraction [21]. It must be stressed that the method involved deconjugation with β-glucuronidase and indirect and non-specific determination of conjugate forms by LC-MS/MS: the differences in the sample preparation and analysis only permitted a qualitative comparison.…”

Section: Analytementioning

confidence: 99%

Determination of α- and β-boldenone sulfate, glucuronide and free forms, and androstadienedione in bovine urine using immunoaffinity columns clean-up and liquid chromatography tandem mass spectrometry analysis

Chiesa

Pavlović

Dusi

et al. 2015

Talanta

Self Cite

View full text Add to dashboard Cite

Section: Analytementioning

confidence: 99%

Determination of α- and β-boldenone sulfate, glucuronide and free forms, and androstadienedione in bovine urine using immunoaffinity columns clean-up and liquid chromatography tandem mass spectrometry analysis

Chiesa

Pavlović

Dusi

et al. 2015

Talanta

Self Cite

View full text Add to dashboard Cite

“…In spite of the excellence in applying all kinds of steroids, this method requires a lengthy procedure (2.5 d) and plenty of poisonous chemical reagents (900 ml per sample). To overcome the shortcomings, some authors used simple SPE methods to take the place of column chromatographic method (Draisci et al, 2003), coupled SPE to HPLC (van Poucke and van Peteghem, 2002), and employed the liquid-liquid extraction step (Gasparini et al, 2007), immunoaffinity columns (Barrón et al, 1996), or solid-phase microextraction (SPME) (Saito et al, 2010). These observations led us to consider an alternative extraction procedure which would be less timeconsuming and more suitable for routine analysis.…”

Section: Extraction and Purification Of Steroidsmentioning

confidence: 99%

Analysis of 19-nortestosterone residue in animal tissues by ion-trap gas chromatography-tandem mass spectrometry

Jiang

Zhang

et al. 2011

J. Zhejiang Univ. Sci. B

View full text Add to dashboard Cite

A rapid sample treatment procedure for the gas chromatography-tandem mass spectrometry (GC-MS) determination of 19-nortestosterone (19-NT) in animal tissues has been developed. In our optimized procedures, enzymatic hydrolysis with β-glucuronidase from Escherichia coli was performed in an acetate buffer (pH 5.2, 0.2 mol/L). Next, the homogenate was mixed with methanol and heated at 60 °C for 15 min, then placed in an ice-bath at −18 °C for 2 h. After liquid-liquid extraction with n-hexane, the analytes were subjected to a normal-phase solid phase extraction (SPE) C 18 cartridge for clean-up. The dried organic extracts were derivatized with heptafluorobutyric anhydride (HFBA), and then the products were injected into GC-MS. Using electron impact mass spectrometry (EI-MS) with positive chemical ionization (PCI), four diagnostic ions (m/z 666, 453, 318, and 306) were determined. A standard calibration curve over the concentration range of 1-20 ng/g was reached, with Y=467 084X−68 354 (R 2 =0.999 7) for 19-NT, and the detection limit was 0.3 ng. When applied to spiked samples collected from bovine and ovine, the recoveries ranged from 63% to 101% with relative standard deviation (RSD) between 2.7% and 8.9%. The procedure is a highly efficient, sensitive, and more economical method which offers considerable potential to resolve cases of suspected nandrolone doping in husbandry animals.

show abstract

“…6 Currently, several methods exist to differentiate between free and conjugated boldenone metabolites but most of these methods have shortcomings. One of these methods cannot differentiate between the glucuronide and the sulphate fractions, 21 another is only able to determine the conjugated status of a-and b-boldenone, 22 and most can only detect one of the reported potential alternative markers. [21][22][23] The aim of this work was the development of an analytical method that would allow the separation, detection and quantification of several potential alternative markers of b-boldenone abuse and to determine their conjugated status.…”

mentioning

confidence: 96%

“…One of these methods cannot differentiate between the glucuronide and the sulphate fractions, 21 another is only able to determine the conjugated status of a-and b-boldenone, 22 and most can only detect one of the reported potential alternative markers. [21][22][23] The aim of this work was the development of an analytical method that would allow the separation, detection and quantification of several potential alternative markers of b-boldenone abuse and to determine their conjugated status. The method could thus then later be used in the routine analyses of suspected boldenone samples or even to help clarify the controversy associated with boldenone detection.…”

mentioning

confidence: 96%

Fractionation of free and conjugated steroids for the detection of boldenone metabolites in calf urine with ultra‐performance liquid chromatography/tandem mass spectrometry

Poucke

Vossel

Peteghem

2008

Rapid Comm Mass Spectrometry

View full text Add to dashboard Cite

For over a decade there has been an intensive debate on the possible natural origin of boldenone (androst-1,4-diene-17beta-ol-3-one, 17beta-boldenone) in calf urine and several alternative markers to discriminate between endogenously formed boldenone and exogenously administered boldenone have been suggested. The currently approved method for proving illegal administration of beta-boldenone(ester) is the detection of beta-boldenone conjugates. In the presented method the sulphate, glucuronide and free fractions are separated from each other during cleanup on a SAX column to be able to determine the conjugated status of the boldenone metabolites. The sulphate and glucuronide fractions are submitted to hydrolysis and all three fractions are further cleaned up on a combination of C18/NH2 solid-phase extraction (SPE) columns. Chromatographic separation of the boldenone metabolites was achieved with a Waters Acquity UPLC instrument using a Sapphire C18 (1.7 microm; 2x50 mm) column within 5 min. Detection of the analytes was achieved by electrospray ionisation tandem mass spectrometry. The decision limits of this method, validated according to Commission Decision 2002/657/EC, were 0.08 ng mL(-1) for androsta-1,4-diene-3,17-dione, 0.13 ng mL(-1) for androst-4-ene-3,17-dione, 0.11 ng mL(-1) for 17alpha-boldenone, 0.07 ng mL(-1) for 17beta-boldenone, 0.24 ng mL(-1) for 5beta-androst-1-en-17beta-ol-3-one and 0.58 ng mL(-1) for 6beta-hydroxy-17beta-boldenone. Because of the fractionation approach used in this method there is no need for conjugated reference standards which often are not available. The disadvantage of needing three analytical runs to determine the conjugated status of each of the metabolites was overcome by using fast chromatography.

show abstract

Development and validation of a liquid chromatography–tandem mass spectrometry method for the separation of conjugated and unconjugated 17α- and 17β-boldenone in urine sample

Cited by 8 publications

References 11 publications

Determination of α- and β-boldenone sulfate, glucuronide and free forms, and androstadienedione in bovine urine using immunoaffinity columns clean-up and liquid chromatography tandem mass spectrometry analysis

Determination of α- and β-boldenone sulfate, glucuronide and free forms, and androstadienedione in bovine urine using immunoaffinity columns clean-up and liquid chromatography tandem mass spectrometry analysis

Analysis of 19-nortestosterone residue in animal tissues by ion-trap gas chromatography-tandem mass spectrometry

Fractionation of free and conjugated steroids for the detection of boldenone metabolites in calf urine with ultra‐performance liquid chromatography/tandem mass spectrometry

Contact Info

Product

Resources

About