Routine high-performance liquid chromatographic determination of urinary unconjugated cortisol using solid-phase extraction and ultraviolet detection

Automated on-line SPE LC–MS/MS method to quantitate 6beta-hydroxycortisol and cortisol in human urine: Use of the 6beta-hydroxycortisol to cortisol ratio as an indicator of CYP3A4 activity

“…Immunochemical results have been shown to overestimate both 6beta-HC and cortisol concentrations due to antibody cross-reactivity with other structurally similar steroids [8]. HPLC methods with colorimetric and fluorometric detection have been shown to give falsely high cortisol and or 6beta-HC values caused by interfering substances with similar retention times as those of cortisol and/or 6beta-HC [17]. Ohno et al addressed most of these problems with their atmospheric pressure chemical ionization (APCI) LC-MS/MS method [18].…”

“…In general, an increase in 6beta-HC:cortisol indicates that the drug may be a potential CYP3A4 inducer and a decrease in the ratio indicates that the drug may be a CYP3A4 inhibitor. Several methods for determinations of 6beta-HC:cortisol in human urine have been published based on enzyme linked immunosorbent assay (ELISA) [8], Radioimmunoassay (RIA) [9], high performance liquid chromatography (HPLC) [10][11][12][13][14][15][16][17] and liquid chromatography-tandem mass spectrometry (LC-MS/MS) [18]. Immunochemical results have been shown to overestimate both 6beta-HC and cortisol concentrations due to antibody cross-reactivity with other structurally similar steroids [8].…”

Improved HPLC Method for the Determination of Cortisol and Cortisone in Urine. Verbesserte HPLC-Methode für die Bestimmung von Cortisol und Cortison im Urin

“…The broader use of HPLC technology for the determination of urinary cortisol and cortisone has been limited by shortcomings of sample extraction, in our experience with previously described methods [5][6][7][8][9][10][11][12], in a large portion of clinical samples quantification is impossible due to impurity of the chromatograms; in most of the methods cited no chromatograms of patients' samples are shown. The main problem in HPLC analysis of urinary cortisol is the large unspecific background produced by phenolic or other interfering substances.…”

“…When applying the simple ultraviolet detection principle for quantification of these compounds, a highly effective sample clean-up is necessary due to the presence of a large quantity of steroid metabolites in urine. Published methods for the determination of cortisol and cortisone in urine with ultraviolet detection [4][5][6][7][8][9][10][11][12] apply solvent extraction, solidphase extraction or solid-phase extraction followed by . solvent extraction.…”