Stability of Urinary Thromboxane A2 Metabolites and Adaptation of the Extraction Method to Small Urine Volume

Background: Thromboxane (TX) A(2) is a pro-thrombotic prostanoid synthesized by activated platelets, biotransformed into 11-dehydro-TXB2, measurable in urines. Eleven-dehydro-TXB2 excretion is increased in high risk cardiovascular diseases; however, this cardiovascular biomarker awaits validation in large trials. The need of large urine volume (8 - 10 mL) and the unknown stability of 11-dehydro-TXB2 in urine after collection might limit its implementation. Methods: We scaled the original method for urine extraction and 11-dehydro-TXB2 measurement down to 1 mL, and assessed its stability at 4 degrees C or 25 degrees C up to 6 days after collection. The sensitivity of the 1 mL procedure was also tested in aspirin-treated patients with low 11-dehydro-TX.132 excretion. Results: The 1 mL adapted method was highly correlated with the original assay (rho = 0.98, p < 0.001, n = 33). Both methods showed similar recoveries in samples spiked with exogenous 11-dehydro-TXB2. Urinary 11-dehydro-TXB2 values in samples immediately frozen were comparable and highly correlated to values in samples at 4 degrees C (day 6: rho = 0.99, p > 0.001, n = 8) or 25 degrees C (day 6: rho = 0.98, p < 0.001, n = 23) up to 6 days in controls and patients. Conclusions: Eleven-dehydro-TXB2 can be measured in small urine volumes and is relatively stable for a few days after collection, even at 25 degrees C. These data allow the validation of this non-invasive cardiovascular biomarker in large studies.