Emory Integrated Lipidomics Core, Emory University School of Medicine, Atlanta, GA 30322, USA.
Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA.
Biomolecules. 2019 May 23;9(5):200. doi: 10.3390/biom9050200.
Liquid-chromatography mass spectrometry is commonly used to identify and quantify metabolites from biological samples to gain insight into human physiology and pathology. Metabolites and their abundance in biological samples are labile and sensitive to variations in collection conditions, handling and processing. Variations in sample handling could influence metabolite levels in ways not related to biology, ultimately leading to the misinterpretation of results. For example, anticoagulants and preservatives modulate enzyme activity and metabolite oxidization. Temperature may alter both enzymatic and non-enzymatic chemistry. The potential for variation induced by collection conditions is particularly important when samples are collected in remote locations without immediate access to specimen processing. Data are needed regarding the variation introduced by clinical sample collection processes to avoid introducing artifact biases. In this study, we used metabolomics and lipidomics approaches paired with univariate and multivariate statistical analyses to assess the effects of anticoagulant, temperature, and time on healthy human plasma samples collected to provide guidelines on sample collection, handling, and processing for vaccinology. Principal component analyses demonstrated clustering by sample collection procedure and that anticoagulant type had the greatest effect on sample metabolite variation. Lipids such as glycerophospholipids, acylcarnitines, sphingolipids, diacylglycerols, triacylglycerols, and cholesteryl esters are significantly affected by anticoagulant type as are amino acids such as aspartate, histidine, and glutamine. Most plasma metabolites and lipids were unaffected by storage time and temperature. Based on this study, we recommend samples be collected using a single anticoagulant (preferably EDTA) with sample processing at <24 h at 4 °C.
液相色谱-质谱联用通常用于鉴定和定量生物样本中的代谢物，以深入了解人体生理学和病理学。代谢物及其在生物样本中的丰度是不稳定的，容易受到采集条件、处理和操作的变化的影响。样本处理的变化可能会以与生物学无关的方式影响代谢物水平，最终导致结果的错误解释。例如，抗凝剂和防腐剂会调节酶活性和代谢物氧化。温度可能会改变酶促和非酶促化学反应。当样本在没有即时处理样本的情况下在偏远地区采集时，采集条件引起的变化的可能性尤其重要。需要有关临床样本采集过程引起的变化的数据，以避免引入人为偏见。在这项研究中，我们使用代谢组学和脂质组学方法以及单变量和多变量统计分析来评估抗凝剂、温度和时间对健康人血浆样本的影响，为疫苗学提供样本采集、处理和加工的指南。主成分分析表明，样本采集程序聚类，抗凝剂类型对样本代谢物变化的影响最大。甘油磷脂、酰基肉碱、鞘脂、二酰基甘油、三酰基甘油和胆固醇酯等脂质以及天冬氨酸、组氨酸和谷氨酰胺等氨基酸都受到抗凝剂类型的显著影响。大多数血浆代谢物和脂质不受储存时间和温度的影响。基于这项研究，我们建议使用单一抗凝剂（最好是 EDTA）采集样本，并在 4°C 下 <24 小时内进行样本处理。
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