Detection of thyroid hormones T3 and T4 in urine and serum after levothyroxine administration: A pilot study relevant...

Thyroid hormones T3 and T4 are clinically relevant hormones that are currently not included on the World Anti-Doping Agency Prohibited List. Pending their inclusion, the possibility of detecting T4 doping was investigated in a T4 administration study involving six healthy volunteers. This pilot study entailed the administration of 1 mg Levothyroxine and the collection of urine and serum samples pre- and post-administration. T3 and T4 quantification methods employing liquid-liquid extraction and liquid chromatography mass spectrometry were developed and validated to analyse all collected samples. The urine and serum steroid profiles were also determined for all samples. All volunteers showed significantly elevated T4 serum concentrations, at least up to 24 h post-administration. From all investigated detection methods, tracking serum T4 concentrations through an individual passport program, employing a 99.7% confidence interval as thresholds, provided the highest test sensitivity, correctly identifying 75% of all post-administration samples as positive. Tracking the T4/T3 ratio in serum provided an identical test sensitivity but requires the additional quantification of T3. No evidence was found indicating the need to include T4 as a confounding factor for the urine or serum steroid profile. Future research should focus on optimizing the extraction protocol and performing a study where T3 is administered in multiple doses and serum samples are collected for a longer period post-administration.