Determination of the metabolites of gestrinone in human urine by high performance liquid chromatography, liquid chromatography/mass spectrometry and gas chromatography/mass spectrometry

Kim, Yunje; Lee, Yongkwan; Kim, Myung-Soo; Yim, Yong‐Hyeon; Lee, Won

doi:10.1002/1097-0231(20000930)14:18<1717::aid-rcm65>3.0.co;2-2

Cited by 17 publications

(7 citation statements)

References 7 publications

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“…By HPLC/MS in ESI positive ion mode, both the in vivo metabolite and Md‘ have [M + H] + at m / z 325 with a major fragment at m / z 263. HPLC/MS/MS on the ion at m / z 263 led to similar fragmentation patterns with major daughter ions at m / z 235, 217, 205, 167, and 153 . Furthermore, the analysis of its TMS-enol TMS-ether derivative by GC/MS, reported in the literature, led to formation of fragments at m / z 423 ([M − 117 u] +. )…”

Section: Resultssupporting

confidence: 61%

“…and 117 (e.g., [CH(OSi(CH 3 ) 3 )CH 3 ] +. ), characteristic for an ethyl side chain containing a TMS-ether moiety . Based on these observations, the metabolite of gestrinone identified in vitro in human hepatocyte incubations (Md) could correspond to this unidentified urinary metabolite reported in the literature following a per os dose of gestrinone in humans.…”

Section: Resultsmentioning

confidence: 67%

“…When gestrinone was dosed orally in human, parent and urinary metabolites were observed in the hydrolyzed fraction of urine samples . These were identified as glucuronide adducts by enzymatic hydrolysis using β-glucuronidase.…”

Section: Resultsmentioning

confidence: 99%

“…Subsequent biosynthesis, isolation, and characterization by high-pressure liquid chromatography (HPLC) coupled to photodiode array (PDA), fluorescence, mass spectrometer (MS) or MS/MS detectors, and NMR were done to elucidate the structure of the major metabolites. To evaluate the in vivo relevance of these new metabolites of THG, in vitro versus in vivo correlation was assessed for gestrinone based on urinary metabolites previously reported in the literature in humans following per os administration …”

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confidence: 99%

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Discovery, Biosynthesis, and Structure Elucidation of Metabolites of a Doping Agent and a Direct Analogue, Tetrahydrogestrinone and Gestrinone, Using Human Hepatocytes

et al. 2005

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Tetrahydrogestrinone (18a-homo-pregna-4,9,11-trien-17beta-ol-3-one, THG) is an anabolic androgenic steroid sold to athletes as an undetectable performance enhancer. Being an unapproved substance, no legitimate in vivo human excretion studies could be performed to identify urinary markers of this doping agent. In vitro systems were used as an alternative approach to study the human metabolism of THG and the gestrinone analogue, which is a marketed drug. Incubations of both compounds in the presence of human hepatocytes led to formation of oxidative and glucuroconjugated metabolites. Microgram quantities of the major in vitro metabolites were biosynthesized using human hepatocytes, characterized by HPLC/MS/MS, and their structures elucidated by NMR. Due to high structure similarity, both THG and gestrinone had an analogous in vitro metabolic pathway leading to successive addition of a hydroxyl and a beta-glucuronic acid at C-18. This in vitro metabolite of gestrinone was consistent with a previously reported major but unknown human urinary metabolite. The structure of another metabolite of THG was proposed to be a glucuroconjugate of an oxidative product with a hydroxyl group most likely at C-16epsilon. In vitro information reported therein could significantly impact the identification of new urinary markers of THG for doping control purposes.

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Section: Resultssupporting

confidence: 61%

Section: Resultsmentioning

confidence: 67%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Discovery, Biosynthesis, and Structure Elucidation of Metabolites of a Doping Agent and a Direct Analogue, Tetrahydrogestrinone and Gestrinone, Using Human Hepatocytes

et al. 2005

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“…In summary, we have presented a method for the quantitative determination of testosterone to epitestosterone ratios in urine that meets or exceeds WADA specifications for LODs as well as qualitative guidelines for the confirmation of banned substances. This method should contribute to the growing use of LC/MS for the identification of steroids for doping control 16–19. The utilization of GRP derivatization combined with LC/Q‐ToF analysis has a number of advantages over traditional GC/MS analysis, including simpler and easier derivatization, faster analysis speed, increased sensitivity, and higher quality mass spectra for unambiguous qualitative determination.…”

Section: Discussionmentioning

confidence: 99%

Quantitative confirmation of testosterone and epitestosterone in human urine by LC/Q‐ToF mass spectrometry for doping control

2008

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Testosterone (T) is the primary male sex hormone. In addition to the development of secondary sex characteristics, testosterone has anabolic effects including increases in muscle size and strength and increases in lean body mass, making it an attractive candidate to enhance athletic performance. In the case of exogenous administration of testosterone, the ratio of testosterone to its isomer, epitestosterone (E), is elevated. WADA has set a standard for T/E ratios of 4.0 as indicative of possible exogenous testosterone administration. Typically, a sample that screens for a T/E ratio above that threshold is then subjected to quantitative confirmation by GC/MS. This methodology, however, can limited due to sensitivity issues as well as a limited number of qualifying ions that can be used for unambiguous identification. We have developed a confirmation method which uses liquid/liquid extraction, followed by room temperature Girard P derivatization, and analysis using LC/MS-ToF. We observe a number of advantages over conventional GC/MS analysis. Analysis time is decreased. Sensitivity is increased, resulting in limits of detection of 2 and 0.5 ng/ml for testosterone and epitestosterone, respectively. The number of diagnostic qualifier ions is also increased allowing more confident identification of the analytes. Finally, while this method has been developed on a QToF instrument, it should be easily transferable to any tandem LC/MS/MS system.

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Current Awareness

2001

J. Mass Spectrom.

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In order to keep subscribers up‐to‐date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of mass spectrometry. Each bibliography is divided into 11 sections: 1 Books, Reviews & Symposia; 2 Instrumental Techniques & Methods; 3 Gas Phase Ion Chemistry; 4 Biology/Biochemistry: Amino Acids, Peptides & Proteins; Carbohydrates; Lipids; Nucleic Acids; 5 Pharmacology/Toxicology; 6 Natural Products; 7 Analysis of Organic Compounds; 8 Analysis of Inorganics/Organometallics; 9 Surface Analysis; 10 Environmental Analysis; 11 Elemental Analysis. Within each section, articles are listed in alphabetical order with respect to author (4 Weeks journals ‐ Search completed at 1st. Nov. 2000)

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Determination of the metabolites of gestrinone in human urine by high performance liquid chromatography, liquid chromatography/mass spectrometry and gas chromatography/mass spectrometry

Cited by 17 publications

References 7 publications

Discovery, Biosynthesis, and Structure Elucidation of Metabolites of a Doping Agent and a Direct Analogue, Tetrahydrogestrinone and Gestrinone, Using Human Hepatocytes

Discovery, Biosynthesis, and Structure Elucidation of Metabolites of a Doping Agent and a Direct Analogue, Tetrahydrogestrinone and Gestrinone, Using Human Hepatocytes

Quantitative confirmation of testosterone and epitestosterone in human urine by LC/Q‐ToF mass spectrometry for doping control

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