A rapid analytical method for the detection of plasma volume expanders and mannitol based on the urinary saccharides...

A screening procedure specifically developed for the detection of saccharides and polyalcohols in human urine in the framework of doping control analysis is presented. The proposed method, set-up, and validated to detect the abuse of dextran, hydroxyethyl starch and mannitol as a doping practice in sport, involves only one enzymatic hydrolysis step and the direct injection into a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. The chromatographic conditions were optimized to allow the efficient separation of compounds with the same molecular weight. Good linearity (R(2) 0.990-0.995) and reproducibility of relative retention times (CV% lower than 1) and of relative abundances of characteristic ion transitions (CV% lower than 10) were obtained. The lower limits of detection and quantification were in the range of 30-100 µg/ml. Since the analytes studied are present also in non-doping products (e.g. in fruit as well as in food products and drugs additives), the developed method was also used to establish a range of reference urinary concentrations: 600 doping control samples and 30 samples from volunteers not using any medication were considered. While the hydrolysis products (isomaltose and maltose hydroxyl-ethylated), used as specific markers for the detection of dextran and hydroxyethyl starch abuse, were not detected in urine; mannitol was present in all urines in a concentration range of 30-1200 µg/ml. Since no criteria of positivity for mannitol has been established yet, the results obtained in this study could be considered, in combination with those of previous researches, as a starting point to fix a threshold value for doping control purpose.