Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 247
Filtrar
Mais filtros










Filtros aplicados

Base de dados
Intervalo de ano de publicação
1.
J Anal Toxicol ; 44(2): 192-199, 2020 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-31322674

RESUMO

In Europe, chemical castration has been adopted as a treatment for paraphilia since the 1930s. Among the various chemical castration agents, luteinizing hormone-releasing hormone (LHRH) agonists are now used widely because of their effectiveness and safety. In South Korea, a legislation of chemical castration to control the sexual impulses of sexual offenders was enforced in July 2011. Most of these subjects are treated with leuprorelin acetate, an LHRH agonist, for chemical castration. Despite this, there are few studies that address the long-term influence of LHRH agonists on testosterone (T) and epitestosterone (E) levels in chemical castration subjects. In order to analyze the urinary levels of T in chemical castration subjects, whose T levels are extremely low, we developed and validated an analytical method for the detection of both T and E in human urine using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. The urine samples were hydrolyzed, extracted, and analyzed by LC-MS/MS with electrospray ionization in the positive-ion mode. The limits of detection were 0.02 ng/mL and the limits of quantitation were 0.05 ng/mL, which provided great sensitivity. The established method was applied to urine samples from chemical castration subjects and healthy male volunteers. The chemical castration subjects showed significantly lower urinary T levels than the control subjects. In addition, the urinary E levels were also lower in the chemical castration subjects; however, the T/E ratios were constant and did not show a notable decrease because of the simultaneous decrease in both urinary T and E. The urinary T levels and T/E ratio did not exceed the doping control criteria for exogenous T ingestion for any subject. This study shows the trend of urinary T and E levels in long-term treated chemical castration subjects by establishing a highly sensitive LC-MS/MS method, that provides useful information for monitoring chemical castration.


Assuntos
Castração , Epitestosterona/urina , Testosterona/urina , Adulto , Cromatografia Líquida , Doping nos Esportes , Europa (Continente) , Humanos , República da Coreia , Espectrometria de Massas em Tandem
2.
Drug Test Anal ; 11(10): 1566-1571, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31454165

RESUMO

Testosterone doping remains a prevalent and potent form of drug cheating among elite athletes. In men, the urine testosterone (T) to epitestosterone (E) ratio (T/E ratio) can identify administration of exogenous T by its suppression of endogenous T production through strong negative feedback on endogenous T and E production as well as spill over into urine of extra testosterone. However, this mechanism may be partially inoperative in females whose much lower circulating T derives from three sources, none subject to powerful negative T feedback. Hence, additional methods to detect T doping in females are required. In this study we report two cases of elite female athletes who were sanctioned for T doping proven by measurement of serum T using liquid chromatography-mass spectrometry (LC-MS), when serial urine T and T/E ratio in one were not indicative of T doping, and in the other were nullified by incidental genetic inactivation of T glucuronidation through the uridine diphosphoglucuronosyl transferase 2B17 (UGT2B17) deletion genotype-phenotype. These findings indicate the potential for serum T measurement by LC-MS to detect T doping in female athletes, especially if implemented in the Bayesian format of an athlete biological passport.


Assuntos
Epitestosterona/urina , Detecção do Abuso de Substâncias/métodos , Testosterona/urina , Atletas , Cromatografia Líquida/métodos , Doping nos Esportes , Feminino , Humanos , Espectrometria de Massas/métodos , Testosterona/sangue
3.
Drug Test Anal ; 11(8): 1218-1230, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30932347

RESUMO

The introduction of alternative markers to the steroid profile can be an effective approach to improving the screening capabilities for the detection of testosterone (T) misuse. In this work, endogenous steroid sulfates were evaluated as potential markers to detect intramuscular (IM) T administration. Fourteen sulfate metabolites were quantified using mixed-mode solid-phase extraction and analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Urine samples after a single IM injection (100 mg) of T cypionate to six Caucasian and six Asian healthy male volunteers were analyzed. Principal component analysis (PCA) was used to characterize the sample cohort and to obtain the most useful markers for discrimination between pre- and post-administration samples. For Caucasian volunteers, a separation between pre- and post-administration samples was observed in PCA, whereas for Asian volunteers no separation was obtained. Seventeen ratios between sulfate metabolites were selected and further considered. Detection times (DTs) of each marker were evaluated using individual thresholds for each volunteer. The best results were obtained using ratios involving T and epitestosterone (E) sulfates in the denominator. The best marker was the ratio androsterone sulfate/testosterone sulfate (A-S/T-S) which prolonged the DT 1.2-2.1 times in respect to those obtained using T/E ratio in all Caucasian volunteers and 1.3-1.5 times in two Asian volunteers. Other ratios between A-S or etiocholanolone sulfate and E-S, and sulfates of etiocholanolone, dehydroandrosterone or epiandrosterone, and T-S were also found adequate. These ratios improve the DT after IM T administration and their incorporation to complement the current steroid profile is recommended.


Assuntos
Anabolizantes/urina , Androgênios/urina , Epitestosterona/urina , Sulfatos/urina , Testosterona/urina , Anabolizantes/administração & dosagem , Anabolizantes/metabolismo , Androgênios/administração & dosagem , Androgênios/metabolismo , Grupo com Ancestrais do Continente Asiático , Cromatografia Líquida , Doping nos Esportes , Epitestosterona/administração & dosagem , Epitestosterona/metabolismo , Grupo com Ancestrais do Continente Europeu , Humanos , Injeções Intramusculares , Masculino , Detecção do Abuso de Substâncias , Sulfatos/metabolismo , Espectrometria de Massas em Tandem , Testosterona/administração & dosagem , Testosterona/metabolismo
4.
J Steroid Biochem Mol Biol ; 185: 47-56, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30031148

RESUMO

Growth and development of an embryo or fetus during human pregnancy mainly depend on intact hormone biosynthesis and metabolism in maternal amniotic fluid (AF). We investigated the hormonal milieu in AF and developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of 14 sulfated and 6 unconjugated steroids in AF. 65 A F samples (male: female = 35: 30) of mid-gestation ranging from 16th week of gestation to 25th week of gestation were analyzed. Reference data of 20 steroid levels in AF of healthy women were provided. 13 sulfated and 3 unconjugated steroids were for the first time quantified in AF by LC-MS/MS. Highest concentrations were found for pregnenolone sulfate (PregS: mean ±â€¯SD, 8.6 ±â€¯3.7 ng/mL), 17α-hydroxypregnenolone sulfate (17OHPregS: 4.9 ±â€¯2.0 ng/mL), epitestosterone sulfate (eTS: 7.3 ±â€¯3.6 ng/mL), 16α-hydroxydehydroepiandrosterone sulfate (16OH-DHEAS: 21.5 ±â€¯10.7 ng/mL), androsterone sulfate (AnS: 9.2 ±â€¯7.4 ng/mL), estrone sulfate (E1S: 3.0 ±â€¯3.0 ng/mL), estriol 3-sulfate (E3S: 8.1 ±â€¯4.0 ng/mL) and estriol (E3: 1.2 ±â€¯0.4 ng/mL). Only testosterone (T) showed a significant sex difference (p < 0.0001). Correlations between AF steroids mirrored the steroid metabolism of the feto-placental unit, and not only confirmed the classical steroid pathway, but also pointed to a sulfated steroid pathway.


Assuntos
Líquido Amniótico/química , Segundo Trimestre da Gravidez/fisiologia , Esteroides/análise , 17-alfa-Hidroxipregnenolona/análise , Androsterona/análise , Cromatografia Líquida , Desidroepiandrosterona/análise , Epitestosterona/análise , Estriol/análogos & derivados , Estriol/análise , Estrona/análogos & derivados , Estrona/análise , Feminino , Idade Gestacional , Humanos , Masculino , Gravidez , Pregnenolona/análise , Espectrometria de Massas em Tandem
5.
Drug Test Anal ; 10(11-12): 1744-1754, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30315670

RESUMO

The impact of dehydroepiandrosterone (DHEA) administration has been widely studied for anti-doping purposes in men, whereas only a few studies have been performed in women. In the present study, the impact of DHEA on the steroid profile parameters and their carbon isotopic ratios was explored. Eleven healthy young women and 10 healthy young men received two treatments: One with 100 mg/day of DHEA for 28 days and one with a placebo according to a double-blind crossover protocol. Urine and saliva (only in females) samples were collected before and for 72 hours after each short-term treatment. In all female subjects, concentrations of the urinary parameters of the steroid profile were highly impacted by short-term DHEA administration including epitestosterone (E). Gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) analysis was performed and positive results were observed for E in the four female subjects where E concentration was adequate for such analysis, whereas men results remained negative for E. Last, the ability of the Anti-Doping Administration and Management System (ADAMS) software used for the athlete biological passport to identify such doping was assessed. Of the 11 passports generated for female subjects, 10 were automatically classified as an atypical passport finding (ATPF). For the remaining passport with normal status in one woman, the variability of the concentrations prevented the ADAMS software from adjusting individual limits. The most impacted markers in women were T/E and 5αAdiol/E, with a detection window of 36 hours for 5αAdiol/E. In addition, good correlations were observed for DHEA and T concentrations in urine and saliva in females.


Assuntos
Desidroepiandrosterona/administração & dosagem , Cromatografia Gasosa-Espectrometria de Massas/métodos , Saliva/química , Esteroides/análise , Esteroides/urina , Detecção do Abuso de Substâncias/métodos , Adulto , Biomarcadores/análise , Biomarcadores/urina , Isótopos de Carbono/análise , Isótopos de Carbono/urina , Desidroepiandrosterona/análise , Desidroepiandrosterona/urina , Doping nos Esportes , Método Duplo-Cego , Epitestosterona/análise , Epitestosterona/urina , Feminino , Humanos , Masculino , Testosterona/análise , Testosterona/urina , Adulto Jovem
6.
Steroids ; 138: 82-90, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30033342

RESUMO

Conjugates of 17α-substituted testosterone (1 and 2) and 17ß-substituted epitestosterone (3 and 4) with pyropheophorbide a were synthesized. The scheme consisted of synthesis of 17α-hydroxy-3-oxopregn-4-en-21-oic and 17ß-hydroxy-3-oxopregn-4-en-21-oic acids, and their coupling with pyropheophorbide a by means of either ethylene diamine, or 1,5-diamino pentane linkers. Mutual influence of steroidal and macrocyclic fragments in conjugates molecules was dependent on configuration of C17 and length of linker, that was established by analysis of 1H NMR spectra and molecular models of conjugates. Studies of interaction of conjugates with prostate carcinoma cells revealed that their uptake and internalization were independent on the androgen receptor activity, but dependent on the structure of conjugates, decreasing in the following row: 3 > 4 ≥ 1 > 2. Conjugates significantly decreased the LNCaP and PC-3 cells growth at 96 h incubation. Epitestosterone derivatives 3 and 4 also showed superior anti-proliferative activity versus testosterone ones. Conformationally more rigid conjugates 1 and 3, comprising short linkers, were more active than those with long linkers; conjugate 3 was the most potent.


Assuntos
Antineoplásicos/química , Clorofila/análogos & derivados , Epitestosterona/química , Testosterona/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Clorofila/química , Humanos , Masculino , Células PC-3 , Neoplasias da Próstata/metabolismo , Relação Estrutura-Atividade
7.
Drug Test Anal ; 10(10): 1518-1527, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29797687

RESUMO

The use of testosterone and its pro-drugs, such as dehydroepiandrosterone (DHEA), is currently regulated in horseracing by the application of international testosterone thresholds. However, additional steroidomic approaches, such as steroid ratios, to distinguish overall adrenal stimulation from drug administrations and an equine biological passport for longitudinal steroid profiling of individual animals could be advantageous in equine doping testing. Thus, DHEA concentrations and related ratios (testosterone [T] to DHEA and DHEA to epitestosterone [E]) were assessed in the reference population by quantitative analysis of 200 post-race gelding urine samples using liquid chromatography-tandem mass spectrometry. DHEA concentrations ranged between 0.9 and 136.6 ng/mL (mean 12.8 ng/mL), T:DHEA ratios between 0.06 and 1.85 (mean 0.43), and DHEA:E ratios between 0.21 and 13.56 (mean 2.20). Based on the reference population statistical upper limits of 5.4 for T:DHEA ratio and 48.1 for DHEA:E ratio are proposed with a risk of 1 in 10 000 for a normal outlier exceeding the value. Analysis of post-administration urine samples collected following administrations of DHEA, Equi-Bolic® (a mix of DHEA and pregnenolone) and testosterone propionate to geldings showed that the upper limit for T:DHEA ratio was exceeded following testosterone propionate administration and DHEA:E ratio following DHEA administrations and thus these ratios could be used as additional biomarkers when determining the cause of an atypical testosterone concentration. Additionally, DHEA concentrations and ratios can be used as a starting point to establish reference ranges for an equine biological passport.


Assuntos
Desidroepiandrosterona/urina , Cavalos/urina , Detecção do Abuso de Substâncias/métodos , Animais , Cromatografia Líquida/métodos , Doping nos Esportes , Epitestosterona/urina , Limite de Detecção , Masculino , Espectrometria de Massas em Tandem/métodos , Testosterona/urina
8.
Mol Cell Endocrinol ; 461: 112-121, 2018 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-28870779

RESUMO

Epitestosterone is the 17α-epimer of testosterone and has been described as an anti-androgen, since it inhibits the effects produced by testosterone and dihydrotestosterone via the nuclear androgen receptor (nAR). However, epitestosterone also displays an effect which is similar to the non-classical effect of testosterone, depolarizing the membrane potential of Sertoli cells and inducing a rapid Ca2+ uptake. This study aimed to investigate the effects of a treatment with epitestosterone on developmental parameters of immature rats. Animals were chemically castrated by using the gonadotropin-releasing hormone (GnRH) antagonist cetrorelix and then received a replacement of 7 days with epitestosterone or testosterone. Replacement with either epitestosterone or testosterone restored the anogenital distance (AGD) and testicular weight which had been reduced by chemical castration. The immunocontent of nAR and the nAR-immunoreactivity were reduced by epitestosterone treatment in the testis of both castrated and non-castrated animals. Furthermore, testosterone was unable of changing the membrane potential of Sertoli cells through its non-classical action in the group of animals castrated and replaced with epitestosterone. In conclusion, in relation to the level of protein expression of nAR epitestosterone acts as an anti-androgen. However, it acts in the same way as testosterone when genital development parameters are evaluated. Moreover, in castrated rats epitestosterone suppressed the non-classical response of testosterone, changing the pattern of testosterone signalling via a membrane mechanism in Sertoli cells.


Assuntos
Castração , Epitestosterona/farmacologia , Terapia de Reposição Hormonal , Testículo/crescimento & desenvolvimento , Testosterona/farmacologia , Animais , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Hormônio Liberador de Gonadotropina/análogos & derivados , Hormônio Liberador de Gonadotropina/antagonistas & inibidores , Hormônio Liberador de Gonadotropina/metabolismo , Hormônio Liberador de Gonadotropina/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Ratos Wistar , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Túbulos Seminíferos/efeitos dos fármacos , Túbulos Seminíferos/metabolismo , Células de Sertoli/efeitos dos fármacos , Células de Sertoli/metabolismo , Testículo/efeitos dos fármacos
9.
Drug Test Anal ; 10(3): 575-583, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28671321

RESUMO

Endogenous steroid use can increase urinary testosterone/epitestosterone (T/E) values. In addition, ethanol in amounts >0.5 g per kg of body weight (g/kg) can also increase T/E values. However, the effect of smaller doses of ethanol on T/E values is unknown. The influence of 0.2 and 0.4 g/kg of ethanol on baseline T/E values in 20 men and 20 women with low and high baseline T/E values was investigated and correlated with ethyl glucuronide (EtG) and ethyl sulfate (EtS) concentrations. T/E values for 7 of the women were excluded from the study because of undetectable T concentrations or for other reasons. One man and 1 woman with a high T/E baseline value had a significant increase in their T/E value after ingestion of 0.2 g/kg of ethanol. One man and 2 women with a high T/E baseline, and 1 woman with a low T/E baseline had significantly increased T/E values after ingestion of 0.4 g/kg of ethanol. There was wide variability in peak EtG concentrations and a lack of correlation between ethanol dose and EtG concentrations. Interestingly, 1 man and 2 women with increased T/E values following ethanol ingestion had EtG concentrations below the World Anti-Doping Agency (WADA) cut-off of 5000 ng/mL. These findings demonstrate that small amounts of ethanol can elevate T/E values, with women being more susceptible. In addition, consideration should be given to the lowering of the WADA EtG cut-off to detect samples with elevated T/E values from ingestion of low doses of ethanol.


Assuntos
Consumo de Bebidas Alcoólicas/urina , Epitestosterona/urina , Detecção do Abuso de Substâncias/métodos , Testosterona/urina , Adulto , Doping nos Esportes , Feminino , Cromatografia Gasosa-Espectrometria de Massas/métodos , Glucuronatos/urina , Humanos , Limite de Detecção , Masculino , Ésteres do Ácido Sulfúrico/urina , Espectrometria de Massas em Tandem/métodos , Adulto Jovem
10.
Artigo em Inglês | MEDLINE | ID: mdl-28850889

RESUMO

This paper presents the development and validation of a high-resolution full scan (FS) electron impact ionization (EI) gas chromatography coupled to quadrupole Time-of-Flight mass spectrometry (GC/QTOF) platform for screening anabolic androgenic steroids (AAS) in human urine samples. The World Antidoping Agency (WADA) enlists AAS as prohibited doping agents in sports, and our method has been developed to comply with the qualitative specifications of WADA to be applied for the detection of sports antidoping prohibited substances, mainly for AAS. The method also comprises of the quantitative analysis of the WADA's Athlete Biological Passport (ABP) endogenous steroidal parameters. The applied preparation of urine samples includes enzymatic hydrolysis for the cleavage of the Phase II glucuronide conjugates, generic liquid-liquid extraction and trimethylsilyl (TMS) derivatization steps. Tandem mass spectrometry (MS/MS) acquisition was applied on few selected ions to enhance the specificity and sensitivity of GC/TOF signal of few compounds. The full scan high resolution acquisition of analytical signal, for known and unknown TMS derivatives of AAS provides the antidoping system with a new analytical tool for the detection designer drugs and novel metabolites, which prolongs the AAS detection, after electronic data files' reprocessing. The current method is complementary to the respective liquid chromatography coupled to mass spectrometry (LC/MS) methodology widely used to detect prohibited molecules in sport, which cannot be efficiently ionized with atmospheric pressure ionization interface.


Assuntos
Anabolizantes/urina , Doping nos Esportes/prevenção & controle , Cromatografia Gasosa-Espectrometria de Massas/métodos , Androsterona/urina , Criança , Epitestosterona/urina , Feminino , Humanos , Limite de Detecção , Modelos Lineares , Reprodutibilidade dos Testes
11.
J Steroid Biochem Mol Biol ; 165(Pt B): 212-218, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27328448

RESUMO

Testosterone (T) has traditionally been the most commonly reported doping agent by doping control laboratories. The screening of T misuse is performed by the quantification of six endogenous androgenic steroids and the ratio T/E included in the Athlete Biological Passport (ABP). The inclusion of additional metabolites can improve the screening capabilities of ABP. In this study, the potential of 3α-glucuronide-6ß-hydroxyandrosterone (6OH-Andros3G) and 3α-glucuronide-6ß-hydroxyetiocholanolone (6OH-Etio3G) as markers of T oral administration was evaluated. These glucuronides have been shown to be resistant to enzymatic hydrolysis and their quantification by means of liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) was reported as the only way to obtain feasible results. Urine samples were collected from five volunteers before and after the oral administration of 40mg of T undecanoate and were analyzed by a LC-MS/MS method recently developed. Concentration of 6OH-Andros3G and 6OH-Etio3G compounds and those of the glucuronides of T (TG), epitestosterone (EG), androsterone and etiocholanolone were established and different concentration ratios were calculated. The detection windows (DWs) for the T administration obtained by each selected ratio were compared to the one of TG/EG. The results showed that four out of the nine tested markers presented DWs much larger for all volunteers than those obtained by the World Anti-Doping Agency established T/E marker or other alternative markers. The 6OH-Andros3G/EG, 6OH-Etio3G/EG, 6OH-Andros3G/TG and 6OH-Etio3G/TG markers were able to identify the T abuse up to 96h after the administration, extending our detection capability for the misuse up to 84h more than the classic marker. The importance of these markers was also highlighted by their prolonged capacity to detect the T misuse in the case of one volunteer whose TG/EG barely exceeded his individual threshold. As a consequence, the four markers presented in this study seem to have an exceptional potential as biomarkers of T oral administration.


Assuntos
Glucuronídeos/análise , Detecção do Abuso de Substâncias/métodos , Testosterona/análogos & derivados , Administração Oral , Androsterona/análise , Biomarcadores/análise , Cromatografia Líquida de Alta Pressão , Doping nos Esportes/prevenção & controle , Epitestosterona/análise , Etiocolanolona/análise , Cromatografia Gasosa-Espectrometria de Massas , Voluntários Saudáveis , Humanos , Hidrólise , Masculino , Reprodutibilidade dos Testes , Esteroides/análise , Espectrometria de Massas em Tandem , Testosterona/administração & dosagem , Testosterona/análise
12.
Drug Test Anal ; 9(7): 994-1000, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27706926

RESUMO

The UGT2B17 gene deletion polymorphism is known to correlate to urinary concentration of testosterone-glucuronide and hence this genotype exerts a large impact on the testosterone/epitestosterone (T/E) ratio, a biomarker for testosterone doping. The objective of this study was to assess if DNA isolated from athletes' urine samples (n = 713) obtained in routine doping controls could be targeted for genotyping analysis for future integration in the athlete's passport. A control population (n = 21) including both urine and blood DNA was used for genotyping concordance test. Another aim was to study a large group (n = 596) of authentic elite athletes in respect of urinary steroid profile in relation to genetic variation. First we found that the genotype results when using urine-derived DNA did not correlate sufficiently with the genotype obtained from whole blood DNA. Secondly we found males with one or two UGT2B17 alleles had higher T/E (mean 1.63 ± 0.93) than females (mean 1.28 ± 1.08), p˂0.001. Unexpectedly, we found that several male del/del athletes in power sports had a T/E ˃1. If men in power sport exert a different urinary steroid profile needs to be further investigated. The other polymorphisms investigated in the CYP17A1, UGT2B7 and UGT2B15 genes did not show any associations with testosterone and epitestosterone concentrations. Our results show that genotyping using urine samples according to our method is not useful in an anti-doping setting. Instead, it is of importance for the anti-doping test programs to include baseline values in the ABP to minimize any putative impact of genotype. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
DNA/genética , Epitestosterona/urina , Técnicas de Genotipagem/métodos , Glucuronosiltransferase/genética , Antígenos de Histocompatibilidade Menor/genética , Polimorfismo Genético , Detecção do Abuso de Substâncias/métodos , Testosterona/urina , DNA/sangue , DNA/urina , Doping nos Esportes , Feminino , Deleção de Genes , Humanos , Masculino , Esteroide 17-alfa-Hidroxilase/genética , Testosterona/análogos & derivados
13.
Drug Test Anal ; 9(7): 1034-1042, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27758048

RESUMO

Today's doping tests involving longitudinal monitoring of steroid profiles are difficult in women. Women have more complex hormonal fluctuations than men and commonly take drugs such as hormonal contraceptives that are shown to affect biomarkers used in these doping tests. In this study, we followed six women's urinary steroid profile during one menstrual cycle, including both glucuronides and sulfate conjugated fractions. Additionally, we studied what happens to the steroidal module of the Athlete Biological Passport (ABP) after administration of an emergency contraceptive (levonorgestrel, NorLevo®). The study shows that there are large individual variations in all metabolites included in the ABP and that the administration of emergency contraceptives may lead to suspicious steroid profile findings in the ABP. Urinary epitestosterone concentration increased during the menstrual cycle, leading to a decrease in the testosterone/epitestosterone ratio. The ratios followed in the ABP varied widely throughout the menstrual cycle, the coefficient of variation (CV) ranging from 4 to 99%. There was a 3-fold decrease in epitestosterone 24 h post administration of the emergency contraceptive pill and androsterone, etiocholanolone, and 5ß- androstan-3α,17ß-diol concentrations decreased about 2-fold. When analyzed with the ABP software, one of the six women had an atypical profile after taking the emergency contraceptive. Furthermore, we could not find any alterations in excretion routes (i.e., if the metabolites are excreted as glucuronide or sulfate conjugates) during the menstrual cycle or after administration of emergency contraceptive, indicating no direct effect on phase II enzymes. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Anabolizantes/urina , Anticoncepcionais Pós-Coito/urina , Ciclo Menstrual/urina , Esteroides/urina , Detecção do Abuso de Substâncias/métodos , Adulto , Atletas , Cromatografia Líquida de Alta Pressão/métodos , Doping nos Esportes , Epitestosterona/urina , Feminino , Cromatografia Gasosa-Espectrometria de Massas/métodos , Glucuronídeos/urina , Humanos , Pessoa de Meia-Idade , Testosterona/urina
14.
Drug Test Anal ; 9(9): 1328-1336, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27717154

RESUMO

Detection of testosterone and/or its pro-drugs in the gelding is currently regulated by the application of an international threshold for urinary testosterone of 20 ng/mL. The use of steroid ratios may provide a useful supplementary approach to aid in differentiating between the administration of these steroids and unusual physiological conditions that may result in atypically high testosterone concentrations. In the current study, an ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was developed to quantify testosterone (T) and epitestosterone (E). The method was used to analyze 200 post-race urine samples from geldings in order to generate the ratios for the reference population. Following statistical analysis of the data, an upper limit of 5 for T:E ratio in geldings is proposed. Samples collected from 15 geldings with atypical urinary testosterone concentrations (>15 ng/mL) but otherwise normal steroid profile, had T:E ratios within those observed for the reference population. The applicability of an upper T:E ratio to detect an administration was demonstrated by the analysis of a selection of incurred samples from testosterone propionate, dehydroepiandrosterone (DHEA), and a mixture of DHEA and pregnenolone (Equi-Bolic®) administrations. These produced testosterone concentrations above the threshold of 20 ng/mL, but also T:E ratios above the proposed limit of 5. In conclusion, consideration of the T:E ratio appears to be a valuable complementary aid to evaluate whether an atypical testosterone concentration could be caused by a natural biological outlier as opposed to the administration of these steroids. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Líquidos Corporais/química , Desidroepiandrosterona/análise , Doping nos Esportes/estatística & dados numéricos , Epitestosterona/análise , Esteroides/análise , Espectrometria de Massas em Tandem/métodos , Testosterona/análise , Animais , Cromatografia Líquida , Desidroepiandrosterona/urina , Epitestosterona/urina , Cavalos , Humanos , Pró-Fármacos , Esteroides/urina , Detecção do Abuso de Substâncias , Testosterona/urina
15.
Drug Test Anal ; 8(11-12): 1197-1203, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27714988

RESUMO

The laboratory profile of intranasal testosterone gel has not been previously reported from an anti-doping perspective. Because intranasal testosterone gel is newly available as a commercial product, we sought to examine the laboratory parameters following administration of this formulation, with particular attention to anti-doping guidelines. Five healthy and active male subjects were administered testosterone intranasal gel three times daily for four weeks, using a pattern of five consecutive days on, two days off. Urine was collected after each five-day round of drug administration and analyzed using a full steroid screen and isotope ratio mass spectrometry (IRMS). Windows of detection for elevated testosterone/epitestosterone (T/E) and other steroid ratios, World Anti-Doping Agency (WADA) athlete biological passport (ABP) findings, and IRMS results were analyzed in this study. In the 0-24 h window post-administration, 70% of samples were flagged with a suspicious steroid profile and 85% were flagged as atypical passport findings according to the WADA ABP steroid module. In the 24-48 h window, 0% of samples displayed suspicious steroid profiles while 40% resulted in atypical passport findings. IRMS testing confirmed the presence of exogenous testosterone in 90% and 40% of samples in the 0-24 h and 24-48 h windows post-administration, respectively. Additionally, IRMS data were analyzed to determine commonalities in the population changes in δ13 C values of testosterone, androsterone, etiocholanolone, 5αAdiol, and 5ßAdiol. Though no discernible metabolic trend of the route of administration was identified, we discovered that intranasal gel testosterone is detectable using conventional anti-doping tests. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Administração Intranasal/métodos , Androsterona/análise , Biomarcadores/análise , Isótopos de Carbono/química , Epitestosterona/análise , Etiocolanolona/análise , Cromatografia Gasosa-Espectrometria de Massas/métodos , Esteroides/análise , Detecção do Abuso de Substâncias/métodos , Testosterona/administração & dosagem , Androsterona/química , Atletas , Biomarcadores/metabolismo , Doping nos Esportes , Epitestosterona/química , Etiocolanolona/química , Humanos , Espectrometria de Massas , Esteroides/química , Testosterona/química , Fatores de Tempo
16.
Drug Test Anal ; 7(11-12): 1017-24, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26695489

RESUMO

This study investigated the effect of Ramadan on the haematological and steroid module of the Athletes Biological Passport (ABP) of the World Anti-Doping Agency (WADA). Nine healthy physically active subjects were tested in the morning and afternoon for two days before and three days during Ramadan. Sample collection and all analyses were performed according to WADA technical documents. Although there were significant changes in the haemoglobin concentration during Ramadan, especially during the first fasting week, none of the subjects in this study exceeded the individually calculated thresholds of the ABP. No significant effects on testosterone/epitestosterone (T/E) ratio were observed but only the afternoon specific gravity (SG) of the urine was elevated. Thus, when urinary steroid concentrations are required, SG corrections need to be performed. The haematological and the steroid module of the ABP can be reliably applied during Ramadan as the observed changes are only marginal.


Assuntos
Atletas , Doping nos Esportes , Epitestosterona/urina , Jejum , Hemoglobinas/metabolismo , Islamismo , Substâncias para Melhoria do Desempenho , Detecção do Abuso de Substâncias/métodos , Adulto , Biomarcadores/sangue , Biomarcadores/urina , Jejum/sangue , Jejum/urina , Humanos , Masculino , Substâncias para Melhoria do Desempenho/sangue , Substâncias para Melhoria do Desempenho/urina , Projetos Piloto , Valor Preditivo dos Testes , Contagem de Reticulócitos , Gravidade Específica , Fatores de Tempo , Urinálise , Adulto Jovem
17.
J Steroid Biochem Mol Biol ; 152: 101-13, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25960318

RESUMO

Celecoxib has been reported to switch the human SULT2A1-catalyzed sulfonation of 17ß-estradiol (17ß-E2) from the 3- to the 17-position. The effects of celecoxib on the sulfonation of selected steroids catalyzed by human SULT2A1 were assessed through in vitro and in silico studies. Celecoxib inhibited SULT2A1-catalyzed sulfonation of dehydroepiandrosterone (DHEA), androst-5-ene-3ß, 17ß-diol (AD), testosterone (T) and epitestosterone (Epi-T) in a concentration-dependent manner. Low µM concentrations of celecoxib strikingly enhanced the formation of the 17-sulfates of 6-dehydroestradiol (6D-E2), 17ß-dihydroequilenin (17ß-Eqn), 17ß-dihydroequilin (17ß-Eq), and 9-dehydroestradiol (9D-E2) as well as the overall rate of sulfonation. For 6D-E2, 9D-E2 and 17ß-Eqn, celecoxib inhibited 3-sulfonation, however 3-sulfonation of 17ß-Eq was stimulated at celecoxib concentrations below 40 µM. Ligand docking studies in silico suggest that celecoxib binds in the substrate-binding site of SULT2A1 in a manner that prohibits the usual binding of substrates but facilitates, for appropriately shaped substrates, a binding mode that favors 17-sulfonation.


Assuntos
Inibidores de Ciclo-Oxigenase 2/farmacologia , Estradiol/metabolismo , Pirazóis/farmacologia , Sulfonamidas/farmacologia , Sulfotransferases/metabolismo , Androstenodiol/metabolismo , Sítios de Ligação , Celecoxib , Desidroepiandrosterona/metabolismo , Epitestosterona/metabolismo , Equilina/análogos & derivados , Equilina/metabolismo , Humanos , Modelos Moleculares , Simulação de Acoplamento Molecular , Pirazóis/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Sulfonamidas/metabolismo , Sulfotransferases/genética , Testosterona/metabolismo
18.
Steroids ; 93: 32-8, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25449768

RESUMO

The intratesticular testosterone concentration is high during the early postnatal period although the intracellular androgen receptor expression (iAR) is still absent in Sertoli cells (SCs). This study aimed to evaluate the non-classical effects of testosterone and epitestosterone on calcium uptake and the electrophysiological effects of testosterone (1µM) on SCs from rats on postnatal day (pnd) 3 and 4 with lack of expression of the iAR. In addition, crosstalk on the electrophysiological effects of testosterone and epitestosterone with follicle stimulating hormone (FSH) in SCs from 15-day-old rats was evaluated. The isotope (45)Ca(2+) was utilized to evaluate the effects of testosterone and epitestosterone in calcium uptake. The membrane potential of SCs was recorded using a standard single microelectrode technique. No immunoreaction concerning the iAR was observed in SCs on pnd 3 and 4. At this age, both testosterone and epitestosterone increased the (45)Ca(2+) uptake. Testosterone promoted membrane potential depolarization of SCs on pnd 4. FSH application followed by testosterone and epitestosterone reduced the depolarization of the two hormones. Application of epitestosterone 5 min after FSH resulted in a delay of epitestosterone-promoted depolarization. The cell resistance was also reduced. Thus, in SCs from neonatal Wistar rats, both testosterone and epitestosterone act through a non-classical mechanism stimulating calcium uptake in whole testes, and testosterone produces a depolarizing effect on SC membranes. Testosterone and epitestosterone stimulates non-classical actions via a membrane mechanism, which is independent of iAR. FSH and testosterone/epitestosterone affect each other's electrophysiological responses suggesting crosstalk between the intracellular signaling pathways.


Assuntos
Androgênios/farmacologia , Epitestosterona/farmacologia , Células de Sertoli/fisiologia , Animais , Animais Recém-Nascidos , Cálcio/metabolismo , Hormônio Foliculoestimulante/farmacologia , Hormônio Foliculoestimulante/fisiologia , Masculino , Potenciais da Membrana , Ratos Wistar , Células de Sertoli/efeitos dos fármacos , Testículo/citologia , Testículo/efeitos dos fármacos
19.
Artigo em Inglês | MEDLINE | ID: mdl-25156963

RESUMO

This study proposes a new analytical methodology for the determination of trace levels of testosterone (T) and epitestosterone (E) in urine matrices using bar adsorptive microextraction combined with liquid desorption followed by high-performance liquid chromatography with diode array detection (BAµE-LD/HPLC-DAD). The comparison of different sorbent coatings (five activated carbons, one styrene-divinylbenzene, two modified pyrrolidone, one ciano and one n-vinylpyrrolidone polymers) through BAµE showed that the latter phase presented much higher selectivity and capacity offering multiple mechanisms of interaction. Assays using this phase were performed on 25mL of water samples spiked at the 8.0µg/L level, yielded average recoveries of 92.1 and 93.4% for T and E, respectively, under optimized experimental conditions; BAµE (n-vinylpyrrolidone): 16h (1000rpm), pH 5.5; LD: acetonitrile, 30min under sonication treatment. From the developed analytical methodology, suitable detection limits were achieved (0.4µg/L) and good linear dynamic ranges (1.4-16.0µg/L) with remarkable determination coefficients (r(2)>0.9978). By using the standard addition methodology, the application of the present analytical approach on urine samples revealed good sensitivity. The proposed method, which operated under the floating sampling technology, proved to be a suitable sorption-based static microextraction alternative for screening T, E and the T/E ratio in urine samples for doping control purposes. The methodology showed to be easy to implement, demonstrating good reproducibility, sensitivity and robustness, allowing the possibility to choose the most selective sorbent coating according to the compounds of interest.


Assuntos
Fracionamento Químico/métodos , Doping nos Esportes , Avaliação Pré-Clínica de Medicamentos/métodos , Epitestosterona/urina , Testosterona/urina , Adulto , Cromatografia Líquida de Alta Pressão , Epitestosterona/química , Epitestosterona/isolamento & purificação , Humanos , Limite de Detecção , Modelos Lineares , Masculino , Reprodutibilidade dos Testes , Testosterona/química , Testosterona/isolamento & purificação
20.
Anal Bioanal Chem ; 406(18): 4325-35, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24817358

RESUMO

Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. Discovery of novel biomarkers for early HCC from other liver diseases such as cirrhosis is of great clinical benefit. In this study, a novel steroid hormone metabolomic method based on liquid chromatography-mass spectrometry combined with logistic regression analysis was applied to study the steroid hormone disorders and to screen potential urinary steroid hormone biomarkers of early HCC. Thirty-six urinary steroid hormones were detected and quantified in healthy controls, cirrhotic patients, and early HCC patients. Heat map analysis and multivariate statistical analysis suggested severe disorders of steroid hormone network and holistically decreased urinary steroid hormone pattern in cirrhotic and early HCC patients. Logistic regression analysis reveals that a panel of two urinary steroid hormones (epitestosterone and allotetrahydrocortisol) displayed excellent diagnostic capability for distinguishing early HCC from cirrhosis with area under the curve (AUC) = 0.938 of receiver operating characteristic (ROC) analysis. These results help to overcome the disadvantage of lower sensitivity and specificity of alpha-fetoprotein for distinguishing early HCC from cirrhosis. Our work shows that steroid hormone metabolomics is a promising biomarker tool for biomarker study of early HCC.


Assuntos
Corticosteroides/urina , Biomarcadores/urina , Carcinoma Hepatocelular/urina , Hormônios Esteroides Gonadais/urina , Cirrose Hepática/urina , Neoplasias Hepáticas/urina , Adulto , Área Sob a Curva , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/patologia , Estudos de Casos e Controles , Epitestosterona/urina , Humanos , Cirrose Hepática/diagnóstico , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/patologia , Modelos Logísticos , Masculino , Metabolômica/métodos , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisol/urina , alfa-Fetoproteínas/análise
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA