McHugh Cora E, Flott Thomas L, Schooff Casey R, Smiley Zyad, Puskarich Michael A, Myers Daniel D, Younger John G, Jones Alan E, Stringer Kathleen A
NMR Metabolomics Laboratory, Department of Clinical Pharmacy, College of Pharmacy, University of Michigan, Ann Arbor, MI 48109, USA.
Department of Emergency Medicine, University of Minnesota, Minneapolis, MN 55455, USA.
Metabolites. 2018 Dec 14;8(4):93. doi: 10.3390/metabo8040093.
Though blood is an excellent biofluid for metabolomics, proteins and lipids present in blood can interfere with 1d-¹H NMR spectra and disrupt quantification of metabolites. Here, we present effective macromolecule removal strategies for serum and whole blood (WB) samples.
A variety of macromolecule removal strategies were compared in both WB and serum, along with tests of ultrafiltration alone and in combination with precipitation methods.
In healthy human serum, methanol:chloroform:water extraction with ultrafiltration was compared to methanol precipitation with and without ultrafiltration. Methods were tested in healthy pooled human serum, and in serum from patients with sepsis. Effects of long-term storage at -80 °C were tested to explore the impact of macromolecule removal strategy on serum from different conditions. In WB a variety of extraction strategies were tested in two types of WB (from pigs and baboons) to examine the impact of macromolecule removal strategies on different samples.
In healthy human serum methanol precipitation of serum with ultrafiltration was superior, but was similar in recovery and variance to methanol:chloroform:water extraction with ultrafiltration in pooled serum from patients with sepsis. In WB, high quality, quantifiable spectra were obtained with the use of a methanol: chloroform precipitation.
尽管血液是代谢组学研究中一种出色的生物流体，但血液中存在的蛋白质和脂质会干扰一维氢核磁共振（¹H NMR）谱，并影响代谢物的定量分析。在此，我们介绍针对血清和全血（WB）样本的有效大分子去除策略。
在全血和血清中比较了多种大分子去除策略，并对单独超滤以及超滤与沉淀方法结合的情况进行了测试。
在健康人血清中，将超滤的甲醇：氯仿：水萃取法与有无超滤的甲醇沉淀法进行了比较。这些方法在健康人混合血清以及脓毒症患者的血清中进行了测试。测试了在-80°C长期储存的影响，以探究大分子去除策略对不同条件下血清的影响。在全血中，对两种类型的全血（来自猪和狒狒）测试了多种萃取策略，以检查大分子去除策略对不同样本的影响。
在健康人血清中，超滤的血清甲醇沉淀法更优，但在脓毒症患者的混合血清中，其回收率和方差与超滤的甲醇：氯仿：水萃取法相似。在全血中，使用甲醇：氯仿沉淀法可获得高质量、可定量的谱图。
