Immunoaffinity chromatography combined with gas chromatography—negative ion chemical ionisation mass spectrometry for the confirmation of flumethasone abuse in the equine

Stanley, Shawn M.R.; Wilhelmi, Brendan; Rodgers, J.P.; Bertschinger, H.J.

doi:10.1016/0378-4347(93)80225-s

Cited by 25 publications

(8 citation statements)

References 35 publications

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“…Sepharose or Sephacryl) and kits are available to carry out this procedure (e.g. Stanley et al, 1993). The immobilised antibody is then packed into a column and the matrix percolated through it.…”

Section: Immunoaffinity Extractionmentioning

confidence: 99%

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Makin

Honour

Shackleton³

et al. 2010

Steroid Analysis

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Section: Immunoaffinity Extractionmentioning

confidence: 99%

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Makin

Honour

Shackleton³

et al. 2010

Steroid Analysis

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“…3-(Carboxymethoxyimino)-17α-methyl-5α-androstane-16β,17β-diol (16β-hydroxymestanolone (carboxymethyl)oxime) (15). 16␤-Hydroxymestanolone (9) (137 mg, 0.4 mmol) in pyridine (4 mL) was treated as per the general procedure.…”

Section: Synthesis Of 16β-hydroxymestanolonementioning

confidence: 99%

“…In the main, these approaches rely on antibodies produced against antigens derived from the readily available parent drug. As a result, they are only effective in situations where there is minimal biotransformation of the parent substance [9][10][11][12][13][14][15].…”

Section: Introductionmentioning

confidence: 99%

Analysis of anabolic steroids in the horse: Development of a generic ELISA for the screening of 17α-alkyl anabolic steroid metabolites

Hungerford

Sortais

Smart

et al. 2005

The Journal of Steroid Biochemistry and Molecular Biology

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Due to the potential for misuse of a wide range of anabolic steroids in horse racing, a screening test to detect multiple compounds, via a common class of metabolites, would be a valuable forensic tool. An enzyme-linked immunosorbent assay (ELISA) has been developed to detect 17alpha-alkyl anabolic steroid metabolites in equine urine. 16beta-Hydroxymestanolone (16beta,17beta-dihydroxy-17alpha-methyl-5alpha-androstan-3-one) was synthesised in six steps from commercially available epiandrosterone (3beta-hydroxy-5alpha-androstan-17-one). Polyclonal antibodies were raised in sheep, employing mestanolone (17beta-hydroxy-17alpha-methyl-5alpha-androstan-3-one) or 16beta-hydroxymestanolone conjugated to human serum albumin, via a 3-carboxymethyloxime linker, as antigens. Antibody cross-reactivities were determined by assessing the ability of a library of 54 representative steroids to competitively bind the antibodies. Antibodies raised against 16beta-hydroxymestanolone showed excellent cross-reactivities for all of the 16beta,17beta-dihydroxy-17alpha-methyl steroids analysed and an ELISA has been developed to detect these steroid metabolites. Using this 16beta-hydroxymestanolone assay, urine samples from horses administered with stanozolol (17alpha-methyl-pyrazolo[4',3':2,3]-5alpha-androstan-17beta-ol), were analysed raw, following beta-glucuronidase hydrolysis, and following solid-phase extraction (SPE) procedures. The suppressed absorbances observed were consistent with detection of the metabolite 16beta-hydroxystanozolol. Positive screening results were confirmed by comparison with standard LCMS analyses. Antibodies raised against mestanolone were also used to develop an ELISA and this was used to detect metabolites retaining the parent D-ring structure following methandriol (17alpha-methylandrost-5-ene-3beta,17beta-diol) administration. The ELISA methods developed have application as primary screening tools for detection of new and known anabolic steroid metabolites.

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“…Most of the applications are for preconcentration of drugs from urine, plasma, and milk and only very few deal with preconcentration of environmental pollutants from water or aqueous extracts. IAC has been used to preconcentrate propanolol and LSD from urine samples (7), nortestosterone and related compounds from urine samples (6,8), various estrogen steroids and their glucuronide derivatives from urine samples (9,20), albuterol, digoxin, and PTH (parathyroid hormone) from plasma (10,11,12), aflatoxin M t and zearalenone from milk (15,17) and Cortisol and flumethezone from urine and serum (18,19). Environmental applications of IAC were limited to carbofuran (2), atrazine (73), carbendazim (14), and chlortoluron (16).…”

Section: Iac On-line With Chromatographic Detectorsmentioning

confidence: 99%

Immunoaffinity Extraction with On-Line Liquid Chromatography—Mass Spectrometry

Emon

Lopez-Avila

1996

ACS Symposium Series

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Sample preparations are crucial to the success of all analytical methods. However, sample preparation is often labor-intensive and the rate limiting step of the overall analytical scheme. Certain immunochemical procedures can perform difficult separations, making subsequent analysis simpler and ensuring the specificity of the results. Immunoaffinity chromatography (IAC) is based on the ability of antibodies to extract target analytes, even from complex environmental or biological matrices. High-performance liquid chromatography-mass spectrometry (HPLC/MS) can be coupled to IAC for a complete extraction-detection system. Customized IACs for plasma, urine, water, and milk are discussed in this paper. Portable detection methods which may be suitable to site characterization are described, including capillary electrochromatography coupled with laser-induced fluorescence (CEC/LIF) detection.Immunoaffinity chromatography (IAC) uses the specific ability of antibodies to separate an antigen (target analyte) from a complex matrix. The specific antibody is immobilized onto an inert, solid support (e.g., silica), called an immunoaffinity column and functions as a ligand. Thus, when an aqueous sample containing the target analyte is passed through the immunoaffinity column, the immobilized antibody will selectively remove the target analyte(s) from the aqueous solution. The adsorbed analyte is subsequently removed from the immunoaffinity column with a buffer of lower pH or greater ionic strength than the original solution.This approach to sample preparation has several advantages: (a) it can be highly selective due to the antibody selectivity towards the target compound; (b) it can concentrate large amounts of sample, allowing detection of the analyte at levels that might not be possible with certain chromatographic detectors; (c) it is amenable

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Immunoaffinity chromatography combined with gas chromatography—negative ion chemical ionisation mass spectrometry for the confirmation of flumethasone abuse in the equine

Cited by 25 publications

References 35 publications

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Analysis of anabolic steroids in the horse: Development of a generic ELISA for the screening of 17α-alkyl anabolic steroid metabolites

Immunoaffinity Extraction with On-Line Liquid Chromatography—Mass Spectrometry

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