Simple and sensitive high-performance liquid chromatography method for simultaneous determination of urinary free cortisol and 6β-hydroxycortisol in routine practice: For CYP 3A4 activity evaluation in basal conditions and after grapefruit juice intake

Cytochrome P450 3A4 activity displays a wide variability. The urinary 6β-hydroxycortisol to cortisol ratio, as a non-invasive assay, can be useful for its pretherapeutic characterization. We developed an HPLC–UV method preceded by liquid–liquid extraction for assessment of this ratio in clinical practice. Urine was collected on second void morning-spot sample. Percentage recoveries were high and reproducible. The 6β-hydroxycortisol to cortisol ratio ranged from 1.6 to 9.9 in 12 Caucasian healthy volunteers. It was reduced by 30 to 70% after ingestion of white grapefruit juice, a CYP3A4 inhibitor. Our method, simple, sensitive and accurate, could be helpful for determination of CYP 3A4 activity before oral chemotherapy, or for the monitoring of the use of grapefruit juice as a pharmacological modulator.