Determination and characterization of diuretics in human urine by liquid chromatography coupled to pneumatically assisted electrospray ionization mass spectrometry

Abstract

The aim of this work was to develop a method for the characterization and determination of diuretics in human urine samples by liquid chromatography (LC) coupled to pneumatically assisted electrospray ionization (ES) mass spectrometry (MS). The diuretics studied were substances forbidden by the IOC such as trichlormethiazide, furosemide, canrenoic acid, benzthiazide, bendroflumethiazide, bumetanide, etacrynic acid and spironolactone. For this purpose, the operational parameters of electrospray, such as counter electrode voltage, capillary voltage, sample cone voltage and source temperature, were optimized in order to obtain the best signal stability and the highest sensitivity for the greatest number of diuretic agents. The optimized separation method was successfully coupled with the MS system to analyze the above-mentioned diuretics extracted from spiked urine samples by a liquid extraction and clean-up procedure at basic pH, using ethyl acetate as solvent and the salting-out effect (NaCl). The mass spectra obtained provide adequate information for identification purposes. Positive urine samples obtained from athletes were also analyzed. The presence of these substances in human urine was confirmed by this method, making LC/ES-MS an analytical tool to be considered in the area of antidoping control.

Mass spectrometry in sports drug testing: Structure characterization and analytical assays

“…Drugs belonging to the classes of diuretics, masking agents such as probenecid, and other polar compounds such as RSR 13 (Efaproxiral) are also detectable in comprehensive LC-MS(/MS) screening procedures, in particular after LLE at different pH values or SPE (Sanz-Nebot et al, 2001;Thieme et al, 2001;Deventer et al, 2002;Qin et al, 2003). The heterogeneous nature of diuretics has required sophisticated sample preparation procedures that allow the extraction of a huge variety of analytes, and ESI or APCI have demonstrated sufficient ionization efficiencies in positive and negative ionization modes for alkaline and acidic analytes, respectively.…”

Screening for 18 diuretics and probenecid in doping analysis by liquid chromatography–tandem mass spectrometry

“…LC/MS, however, allows for the direct separation and identification without a preliminary derivatization step. Recently several methods have been reported for the identification of diuretics by LC/MS (n) (Garbis et al, 1998;Sanz-Nebot et al, 2001;Thieme et al, 2001). The aim of this study was to validate a screening method for 18 diuretics and probenecid in human urine following Eurachem validation guidelines (Eurachem Working Group, 1998).…”

Development and validation of a dried blood spot—LC–APCI–MS assay for estimation of canrenone in paediatric samples

“…Several HPLC methods have been reported for estimation of CAN in various biomatrices such as serum, plasma and urine [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26], however there are no published methods which have employed whole blood in liquid form or in the form of DBS. Many of the methods have several limitations such as lack of selectivity [10,11], low sensitivity [13,17] or large volume of biomatrix required (1 ml plasma) [18,25] and as such are not suitable for estimation of CAN in very low volume paediatric blood samples.…”