Detection of testosterone, nandrolone and precursors in horse hair.
Abstract: Growing interest among several horse-breeder associations has initiated the development of a screening procedure to test for anabolic agents in hair, which has the advantage over blood and urine specimens of allowing long-term detection. An analytical method was established to monitor in tails or manes several anabolic substances available as veterinary medicines or as so-called nutritional supplements (clenbuterol, different esters or prohormones of nandrolone and testosterone). The analytical procedure to detect steroids in hair samples consists of the following steps: decontamination of the hair strand or segment with methanol/water (1:1), milling, extraction of the hair material in an ultrasonic bath using methanol, purification by liquid-liquid extraction (n-pentane/methanol, 25:1) and HPLC cleanup, derivatisation of the relevant LC fractions with MSTFA, and measurement using GC-MS/MS technique. The first objective of our study was the detection of exogenous nandrolone (nortestosterone, NT) in the horse hair; therefore nandrolone-associated compounds [nandrolone dodecanoate administered intramuscularly (i.m.) and a mixture of 4-estrenediol and 4-estrenedione, transdermal] were administered to four geldings. The highest concentrations of NT following i.m. treatment were measured after 10 days in a 2-cm hair segment (up to 18 pg/mg); NT was detectable for up to 120 days and in some cases up to 330 days in tail hair (limit of detection 0.3 pg/mg). Following transdermal application, nandrolone as well as the administered prohormones were identified in tail and mane until the latest sampling at 3 months. Furthermore, untreated stallions (128) were investigated to estimate the range of endogenous levels of NT and testosterone (T) in hair. Maximum values of 3 pg/mg (NT) and 1 pg/mg (T) were quantified originating from endogenous formation in the male horse. Additionally, a possible relationship between steroid concentrations in hair specimens and the age of stallions was appraised. NT and T were not detected in hair samples of control geldings. Following nandrolone treatment of geldings, highest values in hair exceeded the endogenous amount detected in untreated stallions. Therefore comparison of concentrations measured in control samples with the estimated endogenous levels could give a clue to exogenous application in cases of abnormally high amounts of NT or T. The possibility of the evaluation of threshold values is discussed as a means to verify an exogenous administration of NT and T in hair samples. Furthermore, the detection of a synthetic substance in hair, e. g. the parent steroid ester by itself, would be unequivocal proof of an exogenous origin of NT or T and the previous medication of the stallion.
Publication Date: 2005-11-09 PubMed ID: 16261327DOI: 10.1007/s00216-005-0104-8Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research aimed to develop a method for detecting anabolic substances, such as testosterone, nandrolone, and their precursors, in horse hair. The method provides longer-term detection than blood and urine tests. The study also investigated normal levels of these substances in untreated stallions and the possibility of using ratio comparison for identifying abnormal concentrations suggestive of exogenous administration of these substances.
Method Development
- The researchers developed an analytical method to detect various anabolic substances in horse hair, including those found in veterinary medicines and nutritional supplements.
- The method involves decontaminating, milling, and extracting the hair samples before a process of purification and derivatisation. The samples are then measured using a GC-MS/MS technique.
Exogenous Application Testing
- Exogenous nandrolone was tested on four geldings (castrated male horses), who were administered with nandrolone-associated compounds intramuscularly or transdermally.
- The highest concentrations of nandrolone following intramuscular treatment were measured after 10 days in a 2-cm hair segment, and the substance was detectable for 120 to 330 days in tail hair.
- After transdermal application, nandrolone and the administered prohormones were detected in tail and mane hair for up to 3 months.
Endogenous Level Estimation
- Untreated stallions were studied to estimate the natural levels of nandrolone and testosterone in horse hair. Maximum values of 3 pg/mg for nandrolone and 1 pg/mg for testosterone were recorded, supposedly formed endogenously in the male horse.
- The research also investigated the relationship between steroid concentrations in hair samples and the age of the stallions, although no specific findings in this area are mentioned.
Nandrolone Treatment of Geldings
- Control geldings showed no detectable levels of nandrolone or testosterone in their hair samples. However, in geldings treated with nandrolone, hair sample concentrations exceeded the endogenous levels recorded in untreated stallions.
Implications and Threshold Values
- The findings suggest that comparing concentrations from control samples with estimated endogenous levels may provide clues to exogenous application of these substances if abnormally high amounts are found.
- Additionally, the possibility of setting threshold values was discussed to verify exogenous administration of these substances.
- The detection of a synthetic compound in hair, such as the parent steroid ester, would be conclusive proof of an exogenous origin.
Cite This Article
APA
Anielski P, Thieme D, Schlupp A, Grosse J, Ellendorff F, Mueller RK.
(2005).
Detection of testosterone, nandrolone and precursors in horse hair.
Anal Bioanal Chem, 383(6), 903-908.
https://doi.org/10.1007/s00216-005-0104-8 Publication
Researcher Affiliations
- Institute of Doping Analysis and Sports Biochemistry, Dresdner Str. 12, 01731 Kreischa, Germany. p.anielski@idas-kreischa.de
MeSH Terms
- Administration, Cutaneous
- Age Factors
- Animals
- Hair / chemistry
- Horses
- Injections, Intramuscular
- Male
- Nandrolone / administration & dosage
- Nandrolone / analogs & derivatives
- Nandrolone / analysis
- Sex Factors
- Tail / chemistry
- Testosterone / analogs & derivatives
- Testosterone / analysis
Citations
This article has been cited 3 times.- Madry MM, Spycher BS, Kupper J, Fuerst A, Baumgartner MR, Kraemer T, Naegeli H. Long-term monitoring of opioid, sedative and anti-inflammatory drugs in horse hair using a selective and sensitive LC-MS/MS procedure.. BMC Vet Res 2016 Jun 1;12:84.
- Stolker AA, Groot MJ, Lasaroms JJ, Nijrolder AW, Blokland MH, Riedmaier I, Becker C, Meyer HH, Nielen MW. Detectability of testosterone esters and estradiol benzoate in bovine hair and plasma following pour-on treatment.. Anal Bioanal Chem 2009 Oct;395(4):1075-87.
- Reiter M, Pfaffl MW, Schönfelder M, Meyer HH. Gene expression in hair follicle dermal papilla cells after treatment with stanozolol.. Biomark Insights 2008 Dec 23;4:1-8.
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