title: High-throughput screening for various classes of doping agents using a new 'dilute-and-shoot' liquid chromatography-tandem mass spectrometry multi-target approach.
abstract: A new multi-target approach based on liquid chromatography--electrospray ionization tandem mass spectrometry (LC-(ESI)-MS/MS) is presented to screen for various classes of prohibited substances using direct injection of urine specimens. With a highly sensitive new generation hybrid mass spectrometer classic groups of drugs--for example, diuretics, beta2-agonists--stimulants and narcotics are detectable at concentration levels far below the required limits. Additionally, more challenging and various new target compounds could be implemented. Model compounds of stimulant conjugates were studied to investigate a possible screening without complex sample preparation. As a main achievement, the integration of the plasma volume expanders dextran and hydroxyethyl starch (HES), commonly analyzed in time-consuming, stand-alone procedures, is accomplished. To screen for relatively new prohibited compounds, a common metabolite of the selective androgen receptor modulator (SARMs) andarine, a metabolite of growth hormone releasing peptide (GHRP-2), and 5-amino-4-imidazolecarboxyamide ribonucleoside (AICAR) are analyzed. Following a completely new approach, conjugates of di(2-ethylhexyl) phthalate (DEHP) metabolites are monitored to detect abnormally high levels of plasticizers indicating for illicit blood transfusion. The assay was fully validated for qualitative purposes considering the parameters specificity, intra- (3.2-16.6%) and inter-day precision (0.4-19.9%) at low, medium and high concentration, robustness, limit of detection (1-70 ng/ml, dextran: 30 µg/ml, HES: 10 µg/ml) and ion suppression/enhancement effects. The analyses of post-administration and routine doping control samples demonstrates the applicability of the method for sports drug testing. This straightforward and reliable approach accomplishes the combination of different screening procedures resulting in a high-throughput method that increases the efficiency of the labs daily work.
authors.affiliation: Institute of Biochemistry and Center for Preventive Doping Research, German Sport University Cologne, Cologne, Germany. s.guddat@biochem.dshs-koeln.de
authors.forename: S
authors.initials: S
authors.lastname: Guddat
authors.forename: E
authors.initials: E
authors.lastname: Solymos
authors.forename: A
authors.initials: A
authors.lastname: Orlovius
authors.forename: A
authors.initials: A
authors.lastname: Thomas
authors.forename: G
authors.initials: G
authors.lastname: Sigmund
authors.forename: H
authors.initials: H
authors.lastname: Geyer
authors.forename: M
authors.initials: M
authors.lastname: Thevis
authors.forename: W
authors.initials: W
authors.lastname: Schänzer
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000652:urine; D000328:Adult; D000368:Aged; D000697:Central Nervous System Stimulants / Q000652:urine; D002853:Chromatography, Liquid / Q000379:methods; D004232:Diuretics / Q000652:urine; D004300:Doping in Sports; D005260:Female; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009294:Narcotics / Q000652:urine; D004364:Pharmaceutical Preparations / Q000652:urine*; D010952:Plasma Substitutes / Q000032:analysis; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D015813:Substance Abuse Detection / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D000697:Central Nervous System Stimulants; D004232:Diuretics; D009294:Narcotics; D004364:Pharmaceutical Preparations; D010952:Plasma Substitutes