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Improved oxytocin analysis from human serum and urine by orbitrap ESI-LC-HRAM-MS.
Drug Testing and Analysis ( IF 2.6 ) Pub Date : 2020-03-11 , DOI: 10.1002/dta.2783 Adrian A Franke 1 , Xingnan Li 1 , Chester Dabalos 2 , Jennifer F Lai 1
Drug Testing and Analysis ( IF 2.6 ) Pub Date : 2020-03-11 , DOI: 10.1002/dta.2783 Adrian A Franke 1 , Xingnan Li 1 , Chester Dabalos 2 , Jennifer F Lai 1
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Native circulating oxytocin (OT) levels in non‐pregnant/non‐lactating/non‐medicated humans are very low (≤ 8 pg/mL). The lower limit of detection (LLOD) of our previous liquid chromatography mass spectrometry (LC–MS) method (10–25 pg/mL) precluded their quantification in serum and urine. Thus, we sought to improve the LC–MS sensitivity of OT measurements in these matrices by hydrophobic tagging and solid phase extraction (SPE). In the former approach, OT was reduced then alkylated with N‐alkyl acetamide (C12, C14, C16, and C18) tags or derivatized using sulfonyl chloride‐based reagents. In the latter approach, native OT in serum and urine was concentrated by offline SPE using gradient acetonitrile washings after first crashing with acetonitrile. Peak urinary eluate fractions were further concentrated online then analyzed by orbitrap‐based LC–MS with electrospray ionization. All hydrophobic OT derivatives had lower sensitivity than native OT. Washing with a water‐acetonitrile gradient during SPE improved the LLOD of OT in spiked serum to 2.5 pg/mL, while adding a subsequent online‐concentration step improved the LLOD in spiked urine to 1–5 pg/mL and allowed us to detect OT in urine from lactating women. We were unable to improve the sensitivity of OT measurements by hydrophobic tagging or by derivatization using sulfonyl chloride‐based reagents. However, we were successful in improving the sensitivity of native OT measurements in serum and urine 2‐ and 5‐fold, respectively, from our previous orbitrap‐based LC–MS method. Offline SPE was mandatory for both matrices and a subsequent online‐concentration step was required for urine.
中文翻译：
改进的Orbitrap ESI-LC-HRAM-MS从人血清和尿中催产素的分析。
非妊娠/非哺乳/非药物治疗的人体内的天然循环催产素（OT）水平非常低（≤8 pg / mL）。我们以前的液相色谱质谱法（LC-MS）的检测下限（LLOD）（10-25 pg / mL）使其无法在血清和尿液中进行定量。因此，我们试图通过疏水标记和固相萃取（SPE）来提高这些基质中OT测量的LC-MS灵敏度。在前一种方法中，先还原OT，然后用N-烷基乙酰胺（C12，C14，C16和C18）标签烷基化，或使用基于磺酰氯的试剂衍生化。在后一种方法中，首先用乙腈速溶后，使用梯度乙腈洗涤液通过离线SPE浓缩血清和尿液中的天然OT。尿液洗脱峰的最高峰进一步在线浓缩，然后通过基于Orbitrap的LC-MS进行电喷雾电离分析。所有疏水性OT衍生物的敏感性均低于天然OT。在SPE期间用水乙腈梯度洗涤将加标血清中OT的LLOD提高到2.5 pg / mL，同时添加后续的在线浓缩步骤将加标尿液中的LLOD提高到1-5 pg / mL，使我们能够检测OT在哺乳期妇女的尿液中。我们无法通过疏水性标记或使用基于磺酰氯的试剂进行衍生化来提高OT测量的灵敏度。但是，我们成功地从以前的基于orbitrap的LC-MS方法中，提高了血清和尿液中天然OT测量的2倍和5倍灵敏度。
更新日期：2020-03-11
中文翻译：
改进的Orbitrap ESI-LC-HRAM-MS从人血清和尿中催产素的分析。
非妊娠/非哺乳/非药物治疗的人体内的天然循环催产素（OT）水平非常低（≤8 pg / mL）。我们以前的液相色谱质谱法（LC-MS）的检测下限（LLOD）（10-25 pg / mL）使其无法在血清和尿液中进行定量。因此，我们试图通过疏水标记和固相萃取（SPE）来提高这些基质中OT测量的LC-MS灵敏度。在前一种方法中，先还原OT，然后用N-烷基乙酰胺（C12，C14，C16和C18）标签烷基化，或使用基于磺酰氯的试剂衍生化。在后一种方法中，首先用乙腈速溶后，使用梯度乙腈洗涤液通过离线SPE浓缩血清和尿液中的天然OT。尿液洗脱峰的最高峰进一步在线浓缩，然后通过基于Orbitrap的LC-MS进行电喷雾电离分析。所有疏水性OT衍生物的敏感性均低于天然OT。在SPE期间用水乙腈梯度洗涤将加标血清中OT的LLOD提高到2.5 pg / mL，同时添加后续的在线浓缩步骤将加标尿液中的LLOD提高到1-5 pg / mL，使我们能够检测OT在哺乳期妇女的尿液中。我们无法通过疏水性标记或使用基于磺酰氯的试剂进行衍生化来提高OT测量的灵敏度。但是，我们成功地从以前的基于orbitrap的LC-MS方法中，提高了血清和尿液中天然OT测量的2倍和5倍灵敏度。
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