The use of high-performance hydrophobic-interaction chromatography (HPHIC) on SynChropak 500 propyl columns has been evaluated for the first time in the analysis of estrogen receptors labeled with [ 1 2 5I]iodoestradiol-17β. These receptors were extracted from reproductive tissues with 500 m M phosphate buffer and applied to the stationary phase. Utilizing an inverse phosphate gradient (500 to 10 m M, elution resulted in rapidly excluded components in the void volume followed by a second radioactive peak at 400 m M phosphate. Both peaks appeared to contain specific estrogen-binding components in that steroid association was inhibited by diethylstilbestrol and free ligand was eluted with a different retention time. A great deal of [ 1 2 5 I]iodoestradiol- 17β was retained by the column. Inclusion of acetonitrile (20%) in the mobile phase resulted in the elution of[ 1 2 5I]iodoestradiol-receptor complexes at a different position from free ligand. Distribution of specific estrogen-binding components appeared to be tumor-dependent. These preliminary results indicate that HPHIC may be useful for isolating various isoforms of steroid hormone receptors so that detailed information regarding their intrinsic properties may be ascertained.
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