Combination of automatic HPLC‐RIA method for determination of estrone and estradiol in serum

Abstract

We developed a highly sensitive assay for estrone and 17 β‐estradiol in serum. Estrone and 17 β‐estradiol, obtained by solid‐phase extraction using a Sep pak tC18 cartridge, were purified by high‐performance liquid chromatography (HPLC). Quantitation of estrone and 17 β‐estradiol were carried out by radioimmunoassay. Not insignificantly, this automatic system of extraction and HPLC succeeded in analyzing 80 samples a week. Intra‐assay coefficients of variation (CV) for estrone and 17 β‐estradiol ranged from 19.5 to 28.7%, and from 8.5 to 13.7%, respectively. The minimum detectable dose for estrone and 17β‐estradiol were 1.04 pg/ml and 0.64 pg/ml, respectively. The serum levels of 17 β‐estradiol using our method strongly correlated with those by Gas chromatography mass spectrometry (GC‐MS). The serum levels of estrone and 17 β‐estradiol in 154 peri‐ and postmenopausal women were estimated to be between 15 and 27 pg/ml and between 3.5 and 24.0 pg/ml, respectively, while the serum level of 17 β‐estradiol in postmenopausal women, in particular, was estimated to be from 3.5 to 6.3 pg/ml. For postmenopausal women who suffered from vasomotor symptoms, the mean levels of estrone and 17 β‐estradiol at 12 to 18 hours after treatment with daily 0.625 mg conjugated equine estrogen (CEE) and 2.5 mg medroxyprogesterone acetate (MPA) were 135.0 and 21.3 pg/ml at 12 months, respectively. On the other hand, levels of estrone and 17 β‐estradiol at 12 to 18 hours after treatment with CEE and MPA every other day, were 73.4 and 15.3 pg/ml, respectively. These highly sensitive assays for estrone and 17 β‐estradiol are useful in measuring low levels of estrogen in postmenopausal women, and monitoring estrogen levels in women receiving CEE as hormone replacement therapy. J. Clin. Lab. Anal. 13:266–272, 1999. © 1999 Wiley‐Liss, Inc.