Cortisol binding in human amniotic fluid

Abstract

We have investigated high-affinity protein binding of cortisol in human amniotic fluid, and changes in binding, if any, during pregnancy. Studies were generally performed on diluted (20 to 25 per cent) amniotic fluid stripped of endogenous steroids by exposure to charcoal. After overnight incubation with amniotic fluid at 4° C. and subsequent gel filtration on Sephadex G-100 at 22°, [3H]-cortisol was eluted with the protein peak. Nonradioactive cortisol (100 ng.) displaced [3H]-cortisol to the elution position of free [3H]-cortisol. After incubation with 3 per cent albumin, [3H]-cortisol was eluted in the position of unbound steroid. [3H]-dexamethasone, incubated with amniotic fluid, was not eluted with the protein peak on subsequent gel filtration. [3H]-cortisol binding in amniotic fluid was displaced by unlabeled cortisol and progesterone but not by testosterone or estradiol. [3H]-testosterone binding in amniotic fluid was displaced by unlabeled testosterone and estradiol, but not cortisol or progesterone. [3H]-cortisol binding was heat stable to 50° C. and not precipitated by 50 per cent (NH4)2SO4. [3H]-testosterone binding was labile at 40° C. and precipitated by 50 per cent (NH4)2SO4. Scatchard analysis of cortisol binding to term amniotic fluid gave a mean Ka of 3.86 ± 0.42 × 108M−1 (S.E.M.) and binding capacity of 577 ng. of cortisol per 100 ml. Similar values were found for amniotic fluid at 14 to 20 weeks. We conclude (1) there is high-affinity, limited-capacity binding of cortisol in human amniotic fluid; (2) the protein-binding characteristics resemble those of corticosteroid binding globulin (CBG) and are distinct from the testosterone-binding globulin.