Quantitative measurement of clenbuterol at the femtomole level in plasma and urine by combined gas chromatography/negative ion chemical ionization mass spectrometry.

A highly sensitive and specific assay was developed for the quantitative measurement of clenbuterol at the femtomole level in human plasma and urine. Clenbuterol and the internal standard (2H9)clenbuterol were measured by gas chromatography/negative ion chemical ionization mass spectrometry with methane as the reagent gas. The two compounds of interest were extracted from the biological samples at pH 13 using ethyl acetate. After two subsequent purification steps, the cleaned-up organic extract was derivatized with pentafluoropropionic anhydride. The mass spectrometer was set to monitor the abundant [M-HCl]- ions of the perfluoroacyl derivatives (m/z 368 and 377), which were generated in the ion source by an electron capture process. This assay required 1 ml of plasma or 0.5 ml of urine and the detection limit of the method was 5 pg ml-1 with a 12.8% relative standard deviation. The accuracy of the clenbuterol assay was also tested day to day with quality control specimens spiked blind to the analyst. The mean difference between the theoretical and actual values was lower than 4.1%.