Pitiranggon Masha, Perzanowski Matthew S, Kinney Patrick L, Xu Dongqun, Chillrud Steven N, Yan Beizhan
Lamont-Doherty Earth Observatory of Columbia University, Palisades, NY, USA Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY, USA.
Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University, New York, NY, USA.
J Chromatogr Sci. 2014 Oct;52(9):1026-32. doi: 10.1093/chromsci/bmt150. Epub 2013 Nov 4.
Exhaled breath condensate (EBC) provides a relatively easy, non-invasive method for measuring biomarkers of inflammation and oxidative stress in the airways. However, the levels of these biomarkers in EBC are influenced, not only by their levels in lung lining fluid but also by the volume of water vapor that also condenses during EBC collection. For this reason, the use of a biomarker of dilution has been recommended. Urea has been proposed and utilized as a promising dilution biomarker due to its even distribution throughout the body and relatively low volatility. Current EBC urea analytical methods either are not sensitive enough, necessitating large volumes of EBC, or are labor intensive, requiring a derivatization step or other pretreatment. We report here a straightforward and reliable LC-MS approach that we developed that does not require derivatization or large sample volume (∼36 µL). An Acclaim mixed-mode hydrophilic interaction chromatography column was selected because it can produce good peak symmetry and efficiently separate urea from other polar and nonpolar compounds. To achieve a high recovery rate, a slow and incomplete evaporation method was used followed by a solvent-phase exchange. Among EBC samples collected from 28 children, urea levels were found to be highly variable, with a relative standard deviation of 234%, suggesting high variability in dilution of the lung lining fluid component of EBC. The limit of detection was found to be 0.036 µg/mL.
呼出气体冷凝物（EBC）为测量气道炎症和氧化应激的生物标志物提供了一种相对简便、无创的方法。然而，EBC中这些生物标志物的水平不仅受其在肺内衬液中的水平影响，还受EBC采集过程中同时冷凝的水蒸气量的影响。因此，有人建议使用稀释生物标志物。由于尿素在全身分布均匀且挥发性相对较低，已被提出并用作一种有前景的稀释生物标志物。目前的EBC尿素分析方法要么灵敏度不够，需要大量的EBC，要么劳动强度大，需要衍生化步骤或其他预处理。我们在此报告一种我们开发的直接且可靠的液相色谱 - 质谱方法，该方法不需要衍生化或大量样品体积（约36 μL）。选择了一款Acclaim混合模式亲水相互作用色谱柱，因为它能产生良好的峰对称性，并能有效地将尿素与其他极性和非极性化合物分离。为了实现高回收率，采用了缓慢且不完全蒸发的方法，随后进行溶剂相交换。在从28名儿童收集的EBC样本中，发现尿素水平变化很大，相对标准偏差为234%，这表明EBC中肺内衬液成分的稀释存在很大差异。检测限为0.036 μg/mL。
