UHPLC-MS/MS bioanalysis of urinary DHEA, cortisone and their hydroxylated metabolites as potential biomarkers for CYP3A-mediated drug–drug interactions

ABSTRACT Aim: A UHPLC-MS/MS assay was developed to quantify urinary dehydroepiandrosterone (DHEA), 7β-hydroxy-DHEA, cortisone and 6β-hydroxycortisone as potential biomarkers to predict CYP3A activity. Results: A sensitive assay at LLOQ of 0.500 ng/ml with good accuracy and precision was developed for the four analytes in human urine. This UHPLC-MS/MS assay was optimized by eliminating nonspecific loss of the analytes in urine, ensuring complete hydrolysis of the conjugates to unconjugated forms and use of the product ions of [M+H-H2O]+ for multiple reaction monitoring detection of DHEA and 7β-hydroxy-DHEA. Conclusion: This assay was successfully applied to a pilot clinical study. It is also...