Turpeinen U, Stenman U H
Helsinki University Central Hospital, Laboratory, Helsinki, Finland.
Scand J Clin Lab Invest. 2003;63(2):143-50.
Measurement of urinary free cortisol is clinically important in the diagnosis of Cushing's syndrome. While liquid chromatography (LC) with UV detection provides much better specificity than immunologic methods, certain drugs cause interference. Detection by mass spectrometry (MS) is a potentially superior method. Our analysis utilizes 1 mL urine spiked with 6-alpha-methylprednisolone as internal standard. The samples were extracted with dichlormethane and the extract was washed, evaporated to dryness and analyzed by LC-MS/MS operating in the negative mode after separation on a reversed-phase C18 column. The calibration curves for analysis of urinary cortisol exhibited consistent linearity and reproducibility in the range of 10-400 nmol/L. Inter-assay CVs were 4.0-7.6%, at mean concentrations of 21-153 nmol/L. The detection limit was 1 nmol/L (signal-to-noise ratio=3). The mean recovery of cortisol added to urine ranged from 67% to 87% and that of the internal standard from 71% to 76%. The regression equation for the LC-MS/MS (x) and HPLC (y) methods was: y=1.095x+8.0 (r=0.996; n=111). Drugs known to interfere with UV detection did not cause problems here. The sensitivity and specificity of the MS/MS method for urinary free cortisol offer advantages over HPLC with UV detection by eliminating drug interference. The higher equipment costs in comparison with HPLC methods using UV detection are balanced by higher throughput, thanks to shorter chromatographic run times.
尿游离皮质醇的测定在库欣综合征的诊断中具有重要的临床意义。虽然带有紫外检测的液相色谱法（LC）比免疫方法具有更好的特异性，但某些药物会产生干扰。质谱法（MS）检测是一种潜在的更优方法。我们的分析使用1 mL加入6-α-甲基泼尼松龙作为内标物的尿液。样品用二氯甲烷萃取，萃取液经洗涤、蒸发至干，然后在反相C18柱上分离后，以负模式通过LC-MS/MS进行分析。尿皮质醇分析的校准曲线在10 - 400 nmol/L范围内呈现出一致的线性和重现性。在平均浓度为21 - 153 nmol/L时，批间变异系数为4.0 - 7.6%。检测限为1 nmol/L（信噪比 = 3）。添加到尿液中的皮质醇平均回收率在67%至87%之间，内标物的平均回收率在71%至76%之间。LC-MS/MS（x）和HPLC（y）方法的回归方程为：y = 1.095x + 8.0（r = 0.996；n = 111）。已知会干扰紫外检测的药物在此处未产生问题。通过消除药物干扰，MS/MS法检测尿游离皮质醇的灵敏度和特异性优于带有紫外检测的HPLC法。由于色谱运行时间较短，通量更高，与使用紫外检测的HPLC方法相比，虽然设备成本较高，但得到了平衡。
