QUANTITATIVE-DETERMINATION BY HPLC OF URINARY 6-BETA-HYDROXYCORTISOL, AN INDICATOR OF ENZYME-INDUCTION BY RIFAMPICIN AND ANTIEPILEPTIC DRUGS

A method for determination of 6 β-hydroxycortisol in urine by means of high performance liquid chromatography is described. After extraction of 10-30 ml aliquots of urine with ethylacetate, separation is accomplished on a silica gel column (30 cm, Lichrosorb Si 100) with a special two-phase four-component eluent of methylene chloride, n-hexane, ethanol and water. Complete separation of α- and β-isomers requires 15 to 20 min. For routine determinations precolumn cleaning by backflush permits injections of samples at minimum time intervals. For quantitative determinations, each injection should contain at least 0.05-0.5 μg of 6 β-hydroxycortisol, depending on the detector employed. The mean excretion rate in healthy male adults (26-40 years) was 273 μg/day (SD=74.5; n=12). In patients on long term mono-therapy with rifampicin, 6 β-hydroxycortisol excretion had risen fourfold (1166 μg/d; SEM=248; n=7), paralleling the known enzyme-inducing effect of rifampicin. The relatively smaller increase to 498 μg/d observed in patients receiving triple therapy with rifampicin, isoniazid and ethambutol points to possible inhibition by isoniazid. The greatest stimulation of 6 β-hydroxycortisol excretion (2352 μg/d) was found in patients receiving antiepileptic therapy (phenytoin and/or carbamazepine and other drugs). The HPLC technique for 6 β-hydroxycortisol proved to be a tool routinely applicable to non-invasive evaluation of drug metabolizing enzyme activity in man. © 1979 Springer-Verlag.