Development of an electrospray ionization mass spectrometric method for the quantification of theophylline in horse serum.
A rapid and selective method has been developed for the determination of theophylline in horse serum by LC-ESI/MS/MS. The analytical method includes a protein precipitation extraction for sample preparation, liquid chromatography separation technique and ionspray tandem mass spectrometry. The drug was extracted from serum using a protein precipitation with acetonitrile and the supernatants were analyzed using an LC-ESI/MS/MS instrument. The chromatography was performed using a 50 x 2.1 mm C(8) analytical column and an isocratic mobile phase composes of 60:40 acetonitrile-0.5% formic acid in water with a flow rate fixed at 350 microL/min. A linear (weighted 1/concentration) relationship was used to perform the calibration over an analytical range of 0.1-20 ppm. The intra-batch precision and accuracy at LLOQ, medium and high concentration were 11.7, 6.9 and 5.4% and 95.8, 107.8 and 95.8%, respectively, and the inter-batch precision and accuracy at LLOQ, medium and high concentration were 10.4, 7.9 and 7.3% and 97.3, 105.2 and 95.9%, respectively. This LC-ESI/MS/MS method for the determination of theophylline in horse serum has been proved to within generally accepted criteria used for bioanalytical assay and was used successfully during clinical investigation.
Beaudry
Francis
F
Department of Veterinary Biomedicine, Faculty of Veterinary Medicine, University of Montreal, St-Hyacinthe, QC, Canada. francis.beaudry@umontreal.ca
Lavoie
Jean-Pierre
JP
Vachon
Pascal
P
D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification; D002138:Calibration; D002853:Chromatography, Liquid; D004300:Doping in Sports / Q000379:methods*; D006736:Horses / Q000097:blood*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods* / Q000592:standards; D013806:Theophylline / Q000097:blood*
D001798:Blood Proteins; D013806:Theophylline