20-and 20-dihydrocortisone may interfere in LC-MS/MS determination of cortisol in saliva and urine
20-and 20-dihydrocortisone may interfere in LC-MS/MS determination of cortisol in saliva and urine
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DOI：
10.1177/0004563217724178
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发表时间：
2018-05-01
影响因子：
2.2
通讯作者：
Brattsand, Goran
中科院分区：
文献类型：
--
作者：
Israelsson, Marlen;Brattsand, Ralph;Brattsand, Goran
LC-MS/MS方法在皮质醇测定中具有高选择性。然而，内源性类固醇代谢物仍可能干扰和损害结果，例如在库欣综合征的诊断中。特别是，在深夜和地塞米松抑制后获得的唾液样本中发现的低浓度皮质醇可能会被错误地升高。方法采用LC-MS/MS法对唾液和尿液中皮质醇的残留时间和质谱与纯候选物质进行比较，鉴定干扰物。为了将目标化合物从干扰物中分离出来，我们的LC-MS/MS方法采用了不同的色谱条件，包括柱和流动相梯度。结果通过调整色谱条件，将原方法中共洗脱和靠近皮质醇洗脱的两种干扰物从皮质醇中分离出来。这些干扰在尿液和唾液中都被发现，并被确定为两种内源性皮质醇异构体20-和20-二氢化可的松。在正离子模式下使用电喷雾电离，同分异构体与皮质醇共享分子质量和质谱碎片模式。两者都产生了m/z 363.1>121.1, 363.1>115.1和363.1>97.1的转变。在我们最初的LC-MS/MS设置中，20-二氢化可的松在色谱步骤中与皮质醇共洗脱，导致假高测定。结论除非将20-二氢化可的松和20-二氢化可的松从皮质醇中分离出来，否则LC-MS/MS测定皮质醇的结果可能会出现错误升高。
Background LC-MS/MS methods offer high selectivity in cortisol determinations. However, endogenous steroid metabolites may still interfere and compromise the results, for example in the diagnosis of Cushing's syndrome. Erroneously elevated cortisol may, in particular, be misleading at the low concentrations found in salivary samples obtained at late night and after dexamethasone suppression.Methods Interferences in our LC-MS/MS method used for determination of cortisol in saliva and urine were identified by comparing their retention times and mass spectra with those of pure candidate substances. The chromatographic conditions used in our LC-MS/MS method, including column and mobile phase gradient, were varied in order to separate the target compound from the interferences.Results Two interferences, which were co-eluting or eluting close to cortisol in our original method, were successfully separated from cortisol by adjustment of the chromatographic conditions. These interferences were found in both urine and saliva and were identified as the two endogenous cortisol isomers 20- and 20-dihydrocortisone. The isomers share molecular mass and mass spectrometric fragmentation pattern with cortisol using electrospray ionization in the positive-ion mode. Both give rise to the transitions m/z 363.1>121.1, 363.1>115.1 and 363.1>97.1. In our original LC-MS/MS setup, the 20-dihydrocortisone co-eluted with cortisol in the chromatography step resulting in false high determinations.Conclusions Cortisol determination by LC-MS/MS may suffer from erroneously elevated results unless 20- and 20-dihydrocortisone are chromatographically separated from cortisol.