Simultaneous determination of testosterone, cortisol, and dehydroepiandrosterone in saliva by stable isotope dilution on-line in-tube solid-phase microextraction coupled with liquid chromatography-tandem mass spectrometry
Simultaneous determination of testosterone, cortisol, and dehydroepiandrosterone in saliva by stable isotope dilution on-line in-tube solid-phase microextraction coupled with liquid chromatography-tandem mass spectrometry
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DOI：
10.1007/s00216-012-6479-4
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发表时间：
2013-01-01
影响因子：
4.3
通讯作者：
Saito, Keita
中科院分区：
文献类型：
--
作者：
Kataoka, Hiroyuki;Ehara, Kentaro;Saito, Keita
建立了一种简便、灵敏的同时测定唾液中睾酮（TES）、皮质醇（CRT）和脱氢表雄酮（DHEA）的方法：采用Discovery HS F5色谱柱，在线固相微萃取（SPME）联用液相色谱-串联质谱（LC-MS/MS）。以Supel-Q PLOT毛细管柱为萃取装置，以200 μ L/min的流速，40 μ L的样品进行25次抽吸/抽出。提取的化合物很容易通过流动相从毛细管中解吸，没有观察到携带。采用稳定同位素标记的内标，对TES、CRT和DHEA的柱状液相色谱-质谱联用方法均具有良好的线性关系，相关系数为0.9998 /千耶和0.9998 /千欧。日内和日内精密度（相对标准偏差）分别低于4.9%和8.5% （n = 5）。该方法可用于分析唾液样品中的TES、CRT和DHEA，且不存在其他预处理峰和干扰峰，TES、CRT和DHEA的定量限（S/N = 10）分别为0.01、0.03和0.29 ng/mL唾液。这些化合物加标到唾液样品中，回收率均在94%以上。该方法用于分析应力和疲劳负荷引起的唾液TES、CRT和脱氢表雄酮水平的变化。
We have developed a simple and sensitive method for the simultaneous determination of testosterone (TES), cortisol (CRT), and dehydroepiandrosterone (DHEA) in saliva by automated online in-tube solid-phase microextraction (SPME) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) using a Discovery HS F5 column. The optimum in-tube SPME conditions were 25 draw/eject cycles of 40 mu L of sample at a flow rate of 200 mu L/min using a Supel-Q PLOT capillary column as an extraction device. The extracted compounds were easily desorbed from the capillary by passage of the mobile phase, and no carryover was observed. The in-tube SPME LC-MS/MS method showed good linearity with correlation coefficients r a parts per thousand yenaEuro parts per thousand 0.9998 for TES, CRT, and DHEA using their respective stable isotope-labeled internal standards. The intra-day and inter-day precisions (relative standard deviations) were below 4.9 and 8.5 % (n = 5), respectively. This method was successfully utilized to analyze TES, CRT, and DHEA in saliva samples without any other pretreatment or interference peaks, and the quantification limits (S/N = 10) of TES, CRT and DHEA were about 0.01, 0.03 and 0.29 ng/mL saliva, respectively. The recoveries of these compounds spiked into saliva samples were each above 94 %. This method was applied to analyze changes in salivary TES, CRT, and DHEA levels resulting from stress and fatigue load.