Accurate and precise cortisol measurements are requisite for ensuring appropriate diagnosis and management of diseases related with adrenal or pituitary gland disorders. Prompted by the needs in characterization of certified reference materials and quality assurance for serum cortisol measurements, we developed and evaluated a highly reliable measurement procedure based on isotope dilution liquid chromatography-tandem mass spectrometry (ID LC-MS/MS) combined with dextran sulfate-Mg2+ precipitation as the sample pretreatment. An appropriate amount of serum was accurately weighed and spiked with the isotope labelled internal standard. After precipitation, massive lipids and lipoproteins were removed from serum matrix. The clear supernatant was transferred and extracted with ethyl acetate-hexane solution. The cortisol was analyzed with LC-MS/MS in positive electrospray ionization mode. The within-run and total coefficient of variations (CVs) ranged from 0.3 to 0.6% and 0.7 to 1.2%, respectively, for a concentration range of 76.30 to 768.04 nmol/L. A regression comparison of the results obtained by the present method and the certified values of ERM-DA451 showed agreement with no statistical difference (Y = 1.0092 X-0.7455; 95% CI for the slope, 0.9940 to 1.0212; 95% CI for the intercept, - 3.6575 to 2.6390, r2 = 0.999). All structural analogs of cortisol tested were well resolved from cortisol in 12 min on a phenyl ligand column under an isocratic elution. The limit of quantification was estimated to 5 pg cortisol in absolute amount. This method is accurate and simple and can be served as a candidate reference measurement procedure in establishment of serum cortisol reference system.

中文翻译：

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同位素稀释液相色谱-串联质谱结合硫酸葡聚糖-Mg2 +沉淀法开发和验证血清皮质醇候选参考方法。

准确和精确的皮质醇测量是确保正确诊断和管理与肾上腺或垂体疾病相关的疾病的必要条件。出于对认证参考材料的表征和血清皮质醇测量质量保证的需求的推动，我们开发并评估了基于同位素稀释液相色谱-串联质谱（ID LC-MS / MS）结合硫酸葡聚糖的高可靠性测量程序。 Mg2 +沉淀作为样品的预处理。准确称取适量的血清，并加同位素标记的内标物。沉淀后，从血清基质中去除大量脂质和脂蛋白。转移透明的上清液，并用乙酸乙酯-己烷溶液萃取。用LC-MS / MS在正电喷雾电离模式下分析皮质醇。对于76.30至768.04 nmol / L的浓度范围，运行内和总变异系数（CV）分别为0.3％至0.6％和0.7％至1.2％。通过本方法获得的结果与ERM-DA451的认证值的回归比较显示无统计学差异的一致性（Y = 1.0092 X-0.7455；斜率的95％CI，0.9940至1.0212；截距的95％CI ，-3.6575至2.6390，r2 = 0.999）。在等度洗脱下，在苯基配体色谱柱上用12分钟的时间从皮质醇中很好地分离了所有皮质醇的结构类似物。定量限估计为绝对量为5 pg皮质醇。