Measuring some sex and precursor steroids is still challenging even by liquid chromatography - tandem mass spectrometry (LC-MS/MS), and few normal values are available. We developed a LC-MS/MS method for estradiol, estrone, dihydrotestosterone and 17-hydroxypregnenolone measurement, compared it with direct immunoassays, and generated sex, age, menopausal and menstrual status specific reference intervals. Liquid-liquid extraction was optimized on 300 μL serum spiked with isotopic internal standards. A 2D-LC system allowed on-line purification and separation in 11 min run. Electrospray ionization was enhanced by ammonium fluoride. MS-detection was obtained by multiple reaction monitoring. Direct ECLIA for estradiol (n = 80) and RIA for estrone (n = 41) were compared with LC-MS/MS. Reference values were estimated in healthy, lean women in reproductive age (n = 118), menopausal women (n = 33) and men (n = 159). The assay showed satisfying imprecision, trueness, recovery and selectivity. Adequate functional sensitivity was achieved for measuring estrone (18.1 pmol/L) and 17-hydroxypregnenolone (117 pmol/L) in all subjects, and estradiol (35.9 pmol/L) and dihydrotestosterone (134 pmol/L) in women in reproductive age and men, but not in menopausal women. Compared with LC-MS/MS, immunoassays showed good agreement for estradiol but severe disagreement for estrone. Estrogens exhibited sex, menopausal and menstrual variations. Dihydrotestosterone and 17-hydroxypregnenolone depended on sex and menopause, the latter also declining with age in men. Strictly defined reference intervals in the adult female and male population were generated for challenging steroids such as estrogens, dihydrotestosterone and 17-hydroxypregnenolone by a novel LC-MS/MS method. Our achievement can be used to deepen the comprehension of several endocrine diseases.

中文翻译：

---

通过液相色谱-串联质谱法对具有挑战性的性激素和前体类固醇的女性和男性血清参考间隔。

即使通过液相色谱-串联质谱（LC-MS / MS），测量某些性别和前体类固醇仍具有挑战性，很少有正常值可用。我们开发了一种用于雌二醇，雌酮，二氢睾丸激素和17-羟基孕烯醇酮的LC-MS / MS方法，并将其与直接免疫测定法进行了比较，并生成了性别，年龄，绝经和月经状态的特定参考间隔。对加有同位素内标物的300μL血清进行了液-液萃取优化。2D-LC系统允许在11分钟内进行在线纯化和分离。氟化铵增强了电喷雾电离。通过多反应监测获得MS检测。将雌二醇的直接ECLIA（n = 80）和雌酮的RIA（n = 41）与LC-MS / MS进行了比较。参考值是在健康，育龄妇女（n = 118），更年期妇女（n = 33）和男性（n = 159）。该测定显示出令人满意的不精确度，真实性，回收率和选择性。在所有年龄和年龄的女性中，测量所有受试者的雌酮（18.1 pmol / L）和17-羟基孕烯醇酮（117 pmol / L）以及雌二醇（35.9 pmol / L）和二氢睾酮（134 pmol / L）时，都具有足够的功能敏感性。男性，但绝经女性则没有。与LC-MS / MS相比，免疫测定显示雌二醇的一致性好，但雌酮的严重分歧。雌激素表现出性别，更年期和月经差异。二氢睾丸激素和17-羟基孕烯醇酮取决于性别和更年期，后者也随着年龄的增长而下降。通过新颖的LC-MS / MS方法，对具有挑战性的类固醇（例如雌激素，二氢睾丸激素和17-羟基孕烯醇酮）生成了成年女性和男性人群中严格定义的参考区间。我们的成就可用于加深对几种内分泌疾病的理解。