Metabolomics profiling of xenobiotics in elite athletes: relevance to supplement consumption

Abstract Background Supplements are widely used among elite athletes to maintain health and improve performance. Despite multiple studies investigating use of dietary supplements by athletes, a comprehensive profiling of serum supplement metabolites in elite athletes is still lacking. This study aims to analyze the presence of various xenobiotics in serum samples from elite athletes of different sports, focusing on metabolites that potentially originate from nutritional supplements. Methods Profiling of xenobiotics in serum samples from 478 elite athletes from different sports (football, athletics, cycling, rugby, swimming, boxing and rowing) was performed using non-targeted metabolomics-based mass spectroscopy combined with ultrahigh-performance liquid chromatography. Multivariate analysis was performed using orthogonal partial least squares discriminant analysis. Differences in metabolic levels among different sport groups were identified by univariate linear models. Results Out of the 102 detected xenobiotics, 21 were significantly different among sport groups including metabolites that potentially prolong exercise tolerance (caffeic acid), carry a nootropic effect (2-pyrrolidinone), exert a potent anti-oxidant effect (eugenol, ferulic acid 4 sulfate, thioproline, retinol), or originate from drugs for different types of injuries (ectoine, quinate). Using Gaussian graphical modelling, a metabolic network that links various sport group-associated xenobiotics was constructed to further understand their metabolic pathways. Conclusions This pilot data provides evidence that athletes from different sports exhibit a distinct xenobiotic profile that may reflect their drug/supplement use, diet and exposure to various chemicals. Because of limitation in the study design, replication studies are warranted to confirm results in independent data sets, aiming ultimately for better assessment of dietary supplement use by athletes

Hyperhydration-induced decrease in urinary luteinizing hormone concentrations of Male athletes in doping control analysis

Low urinary luteinizing hormone (LH) values have been discussed as a marker to detect steroid abuse. However, suppressed LH concentrations related to highly diluted urine samples could be a misleading indication of anabolic steroid abuse. One aim of the present study was to examine the effect of hyperhydration on the interpretation of LH findings during doping control analysis and to investigate different possibilities to correct volume-related changes in urinary LH concentrations. Seven healthy, physically active, nonsmoking White males were examined for a 72-hr period, using water and a commercial sports drink as hyperhydration agents (20 ml/kg body weight). Urine samples were collected and analyzed according to the World Anti-Doping Agency's technical documents. Baseline urinary LH concentrations, expressed as the mean � SD for each individual, were within the acceptable physiological range (7.11 � 5.42 IU/L). A comparison of the measured LH values for both hyperhydration phases (Phase A: 4.24 � 5.60 IU/L and Phase B: 4.74 � 4.72 IU/L) with the baseline (''normal'') values showed significant differences (Phase A: p < .001 and Phase B: p < .001), suggesting the clear effect of urine dilution due to hyperhydration. However, an adjustment of urinary LH concentrations by specific gravity based on a reference value of 1.020 seems to adequately correct the hyperhydration-induced decrease on the LH levels. - 2019 Human Kinetics, Inc.The authors sincerely thank the World Anti-Doping Agency for the financial support of the project (contract 10D21CG). Furthermore, the authors would like to acknowledge the medical staff of Al-Hayat Medical Center, Doha, Qatar, for their help during the clinical part of the project performed under the supervision of Dr. Khaled Youssef, MD and Dr. Fawaz Amin Saad, MD. Finally, Mrs. Noor Al-Motawa, ADLQ Education and Research Office Director, is sincerely acknowledged for her constant support on the research projects. The authors have no conflict of interest to declare.Scopu

Gas chromatographic quadrupole time-of-flight full scan high resolution mass spectrometric screening of human urine in antidoping analysis

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 anti doping 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

High resolution full scan liquid chromatography mass spectrometry comprehensive screening in sports antidoping urine analysis

The aim of this paper is to present the development and validation of a high-resolution full scan (HR-FS) electrospray ionization (ESI) liquid chromatography coupled to quadrupole Orbitrap mass spectrometer (LC/Q/Orbitrap MS) platform for the screening of prohibited substances in human urine according to World Antidoping Agency (WADA) requirements. The method was also validated for quantitative analysis of six endogenous steroids (epitestosterone, testosterone, 5α-dihydrotestosterone, dehydroepiandrosterone, androsterone and etiocholanolone) in their intact sulfates form. The sample preparation comprised a combination of a hydrolyzed urine liquid–liquid extraction and the dilute & shoot addition of original urine in the extracted aliquot. The HR-FS MS acquisition mode with Polarity Switching was applied in combination of the Quadrupole-Orbitrap mass filter. The HR-FS acquisition of analytical signal, for known and unknown small molecules, allows the inclusion of all analytes detectable with LC–MS for antidoping investigations to identify the use of known or novel prohibited substances and metabolites after electronic data files’ reprocessing. The method has been validated to be fit-for-purpose for the antidoping analysis

Additional file 2: Table S1. of A pilot study comparing the metabolic profiles of elite-level athletes from different sporting disciplines

Comparison of previously published metabolite changes in plasma at 60 min after completion of exercise [1] and their corresponding PC2 loading values obtained in this study. Table S2. Metabolites differentiating between moderate- and high-endurance athletes (p ≤ 0.05). Table S3. Metabolites differentiating between moderate- and high-endurance athletes (p ≤ 0.05) in males only. Table S4. Pearson’s Correlations between various sex steroid metabolites. Significant p values are highlighted (* < 0.05, ** < 0.01, *** < 0.001). Table S5. Metabolites differentiating between moderate- and high-power athletes (p ≤ 0.05). Table S6. Metabolites differentiating between moderate- and high-power athletes (p ≤ 0.05) in males only. Table S7. Gender-endurance interaction metabolites. Columns A–F show the effect of endurance on gender-interaction metabolites in males only. Columns H to L show the different effect in females. Table S8. Gender-power interaction metabolites. Columns A–F show the effect of power on gender-interaction metabolites in males only. Columns H to L show the different effect in females. (XLSX 1377 kb

Additional file 1: of A pilot study comparing the metabolic profiles of elite-level athletes from different sporting disciplines

Materials and Methods. (DOCX 16 kb