Dialyzable free cortisol after stimulation with Synacthen®

Objectives To compare the changes in free vs. total serum cortisol concentrations after acute stimulation of the adrenal cortex. Design and methods Paired serum samples of ten individuals taken immediately before and 1 h after stimulation with 250 μg ACTH (1–24) (Synacthen®) given i.v. were analyzed. Total cortisol was quantified using liquid chromatography tandem-mass spectrometry with an online sample extraction system and tri-deuterated cortisol as the internal standard. Free cortisol was measured with the same method after equilibrium dialysis. Concentrations of the corticosteroid-binding globulin (CBG) were determined by radioimmuno assay. Results Total cortisol increased by a mean of 106% (mean basal cortisol 312 nmol/L (SD 140 nmol/L), stimulated 686 nmol/L (SD 163 nmol/L); p< 0.001, paired t-test for differences); no significant change of CBG concentrations was found (874 nmol/L (SD 179 nmol/L) before stimulation, 869 nmol/L (SD 225 nmol/L) after stimulation). The mean increase of free cortisol was 263% (mean basal free cortisol 20.3 nmol/L (SD 13.2 nmol/L), stimulated 73.8 nmol/L (SD 26.7 nmol/L); p< 0.001) and thus substantially more pronounced compared to the increase of total cortisol (p< 0.01). The ratio of free to total serum cortisol was significantly increased after stimulation (6.1% (SD 1.7%) before stimulation, 10.6% (SD 1.9%) after stimulation; p< 0.001). Conclusions After acute neuroendocrine stimulation of the adrenal cortex the relative increase of free bioactive cortisol concentrations is substantially more pronounced than the increase of total cortisol concentrations.