Jackson Thomas C, Zhang Yan Victoria, Sime Patricia J, Phipps Richard P, Kottmann R Matthew
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, United States.
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, United States.
J Chromatogr B Analyt Technol Biomed Life Sci. 2017 Sep 1;1061-1062:468-473. doi: 10.1016/j.jchromb.2017.07.041. Epub 2017 Jul 24.
Exhaled breath condensate (EBC) is easily obtained for clinical diagnosis and prognosis for pulmonary diseases and has gained much interest in biomarker discovery research and studies. Lactate, a physiological material, is found in EBC and has been demonstrated to be a potential indicator of chronic obstructive pulmonary disease and other lung diseases. Several assays are available to detect lactate in human body fluids, and yet none is suitable for detecting lactate in EBC. Due to the very low concentration of lactate in EBC and low volume of EBC, it is very important to develop an assay to measure lactate with high sensitivity, accuracy and easy sample processing. We report here a novel LC-MS/MS based assay to measure lactate using HILIC column separation. Sample preparation was simple and straightforward through a "dilute and shoot" approach with a separation of 4min. The limit of quantification was determined to be 0.5μM. This assay was linear from 0.5μM to at least 100μM. The inter- and intra- day precision at the levels of 1μM, 10μM, and 100μM were less than 3% with recovery within 5.4% of expected values. There was no ion suppression for the assay, and no carry-over was observed up to 500μM. Furthermore, we discovered that lactate is ubiquitously present in the lab environment, which can create significant challenges for accurate detection of lactate at low concentrations. We provided practical approaches in this paper to overcome the challenges and ensure the accuracy of the assay. In summary, this article presents an accurate and sensitive method using LC-MS/MS for measuring lactate in EBC, and this method is suitable for measuring lactate concentrations for non-invasive monitoring of pulmonary functions.
呼出气冷凝物（EBC）易于获取，可用于肺部疾病的临床诊断和预后评估，在生物标志物发现研究中备受关注。乳酸是一种生理物质，存在于EBC中，已被证明是慢性阻塞性肺疾病和其他肺部疾病的潜在指标。有几种检测方法可用于检测人体体液中的乳酸，但没有一种适用于检测EBC中的乳酸。由于EBC中乳酸浓度极低且EBC体积小，开发一种具有高灵敏度、准确性且样品处理简便的检测方法非常重要。我们在此报告一种基于液相色谱-串联质谱（LC-MS/MS）的新型检测方法，使用亲水相互作用色谱（HILIC）柱分离来测量乳酸。样品制备通过“稀释进样”方法简单直接，分离时间为4分钟。定量限确定为0.5μM。该检测方法在0.5μM至至少100μM范围内呈线性。在1μM、10μM和100μM水平下，日内和日间精密度均小于3%，回收率在预期值的5.4%以内。该检测方法不存在离子抑制，在高达500μM时未观察到残留。此外，我们发现实验室环境中普遍存在乳酸，这可能给低浓度乳酸的准确检测带来重大挑战。我们在本文中提供了实际方法来克服这些挑战并确保检测的准确性。总之，本文介绍了一种使用LC-MS/MS准确灵敏地测量EBC中乳酸的方法，该方法适用于测量乳酸浓度以进行肺功能的无创监测。
