High-speed gas chromatography in doping control: Fast-GC and fast-GC/MS determination of β-adrenoceptor ligands and diuretics

Abstract

In official doping controls, about 300 drugs and metabolites have to be screened for each sample. Moreover, the number of determinations to be routinely processed increases continuously as the number of both samples and potential illicit drugs keeps growing. As a consequence, increasingly specific, sensitive, and, above all, fast methods for doping controls are needed. The present study presents an efficient fast-GC/MS approach to the routine screening of two different classes of doping agents, namely beta-adrenoceptor ligands and diuretics (belonging to the S3, P2, and S5 groups of the WADA list of prohibited substances). Narrow bore columns (100 mm id) of different lengths and coated with apolar stationary phases were successfully used to separate the derivatized analytes; preliminary experiments (results not shown) showed better performances with OV-1701 for the separation of beta-adrenoceptor ligands. On the same stationary phase some diuretics required too high a temperature or a long isothermal time for elution, in which case a DB1-MS column was preferred. Two methods of sample preparation, derivatization, and analysis were used on aqueous standard mixtures of, respectively, (i) eight beta-adrenoceptor ligands, including five beta-antagonists (acebutolol, alprenolol, atenolol, metoprolol, pindolol) and three beta2-agonists (salbutamol, clenbuterol, terbutaline) and (ii) seventeen diuretic drugs (acetazolamide, althiazide, bendroflumethiazide, bumethanide, canrenone, chlorothiazide, chlortalidone, clopamide, ethacrinic acid, furosemide, hydrochlorothiazide, hydroflumethiazide, indapamide, indomethacine, spironolactone, triamterene, trichloromethiazide) and one masking agent (probenecid). The mixture of beta-adrenoceptor ligand derivatives was efficiently separated in about 5.6 min, while the one of 18 diuretics and masking agents required less than 5 min for analysis. Limits of detection were from 1 microg/L for pindolol, ethacrinic acid, furosemide, indomethacine, and trichloromethiazide, to 20 microg/L for terbutaline, salbutamol, and metoprolol, and 50 microg/L for clopamide; the instrumental repeatability proved to be excellent (area RSD% <2 for almost all analytes). For this work a quadrupole MS with inert ion source has been used, demonstrating that the quadrupole technology is perfectly adequate to provide precise integration of 400 ms-wide GC peaks.

Chemical Reagents and Derivatization Procedures in Drug Analysis

“…(52) A similar two-step derivatization approach was used for β-adrenoceptor ligands and methyl iodide in potassium carbonate and acetone methylated the diuretics before GC/MS of these mixtures in about 5 min using narrow bore capillary columns. (53) To enhance SPME using a polystyrene-divinylbenzene fiber, phenazone drugs were first acetylated on the nitrogen atom using acetic anhydride but then silylated with MTBSTFA, which removes the acetyl group to enhance GC/MS detection. (54) Pentafluorophenyldimethylsilyl (flophemesyl) derivatives are often generated from steroids by reaction with flophemesylamine at room temperature for 15 min.…”

Determination of 8 Diuretics and Probenecid in Human Urine by Gas Chromatography-Mass Spectrometry: Confirmation Procedure

“…The recovery of diuretics and impeding effect of urine matrix and other diuretics were not studied in this work as well as in [10]. Besides, both methods were not validated.…”

“…Despite the great number of methods proposed, gas or liquid chromatography with mass spectrometric detection do not always provide the necessary selectivity towards the urine components [10,12]. In case of liquid chromatography, the use of tandem mass-spectrometry allowed to pass over these problems and to ensure the satisfactory result [13][14][15].…”

Simultaneous determination of β2‐agonists in human urine by fast‐gas chromatography/mass spectrometry: method validation and clinical application

“…The convenient application of such a technique in antidoping analysis, using short and narrow columns (5-10 m, i.d. 0.05-0.1 mm), fast temperature ramping and high carrier gas fl ow rate was recently demonstrated (Morra et al, 2006;Brunelli et al, 2006;Mazzarino et al, 2007).…”