In vitro synthesis and characterisation of three fenoterol sulfoconjugates detected in fenoterol post-administration urine samples.

Fenoterol, a fast-acting ß2-adrenergic agonist, is used in the therapy of obstructive pulmonary diseases and for the inhibition of premature labour obstetrics. Doping control for ß2-agonists, which are prohibited in sports by the World Anti-Doping Agency, is commonly performed by liquid chromatography/mass spectrometry after hydrolysis of phase II metabolites. The continuing development of analytical procedures has led to direct injection of urine samples without sample preparation becoming a viable tool. For the detection of substances without sample preparation, including hydrolysis, detailed information of the phase II metabolism of the substances is essential. In this study, human S9 fractions of different tissues and two recombinant sulfotransferases were investigated for their potential to form fenoterol sulfoconjugates, which were characterised in detail. Two mono-sulfoconjugates and one bis-sulfoconjugate were synthesised and their structures confirmed by liquid chromatography­high-resolution/high-accuracy mass spectrometry. All of the metabolites were identified as esterified phenolic compounds. Excretion studies with orally and inhalatively administered fenoterol proved the occurrence of the sulfoconjugates in vivo. Inhalatively administered fenoterol resulted in the detection of the two monosulfoconjugates in low amounts in urine due to the lower inhalation dose of fenoterol compared to the oral dose. After oral uptake of fenoterol, the two mono-sulfoconjugates and a fenoterol bis-sulfoconjugate were detected in urine. This is the first report of the bis-sulfoconjugate.

Terbutaline sulfoconjugate: characterization and urinary excretion monitored by LC/ESI-MS/MS.

Terbutaline is a fast-acting beta(2)-adrenergic agonist used in the treatment of obstructive pulmonary diseases. Doping control for beta(2)-agonists, which are forbidden in sports by the World Anti-doping Agency (WADA), is performed in screening by liquid chromatography/mass spectrometry after hydrolysis of phase-II metabolites. In this study, the mono-sulfoconjugated phase-II metabolite of terbutaline was synthesized and the chemical structure was characterized by (1)H-nuclear magnetic resonance spectrometry and high resolution/high accuracy Orbitrap mass spectrometry. The metabolite was designated as the phenolic esterified compound, which has been mentioned in most literature reports but has not been verified so far. The benzylic esterified compound was also synthesized and characterized by high-resolution/high accuracy Orbitrap mass spectrometry but was not detectable in urine samples from an excretion study performed after a single application of one terbutaline capsule (7.5 mg terbutaline sulfate salt). The phenolic sulfate of terbutaline was detected for two to four days after administration, whereas the unchanged terbutaline was detected for four to five days. A glucuronidated, disulfated or trisulfated phase-II metabolite of terbutaline was not found. The measurement of phase-II metabolites is planned to be incorporated into existing screening procedures to allow a faster sample preparation.