IDENTIFICATION OF HUMAN URINARY METABOLITES OF ISBUFYLLINE BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY THERMOSPRAY MASS-SPECTROMETRY

Analysis of urinary metabolites of isbufylline (1, (1, 3-dimethyl-7-(2 -methylpropyl) xanthine) in healthy male volunteers after oral adminis tration of a single dose of 320 mg was undertaken by high-performance liquid chromatography/mass spectrometry (HPLC/MS with thermospray ioni zation. Filtered urines were directed injected into the HPLC/MS system , equipped with a Lichrospher 100 RP 18 analytical column. The mobile phase was 0.1 M, pH 3.7 ammonium acetate in water and acetonitrile; th e composition was varied linearly from 5% to 40% of the organic modifi er in 40 min with a flow rate of 1 ml min(-1). Three more chromatograp hic peaks appeared in urines from treated subjects as compared to untr eated ones; their probable quasi-molecular ions were at m/z 253, 239 a nd 267 respectively, while the original drug, of 236 Da, was not prese nt in appreciable quantity. The collisionally activated daughter ion s pectre of the above ions allowed identification of 1, 3-dimethyl-7-(3- hydroxy-2-methylpropyl) xanthine (D30HMPX), 1-methyl-7-(2-hydroxy-2-me thylpropyl) xanthine (M2OHMPX), and 1, 3-dimethyl-(2-carboxypropyl) xa nthine (D2CMPX), the first one being present as glucuronic acid conjug ate.