开发用于分析哺乳动物生物流体的微流超高效液相色谱-质谱法代谢组学测定。

IF 3.5 3区 医学 Q2 ENDOCRINOLOGY & METABOLISM
Annie J Harwood-Stamper, Caroline A Rowland, Warwick B Dunn
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引用次数: 0

摘要

引言和目的:利用超高效液相色谱-质谱联用技术(UHPLC-MS)进行非靶向代谢组学分析,以研究包括人类在内的生物系统中的新陈代谢的应用正在迅速增加。在其中一些研究中,需要收集和分析低样品量的生物流体(如泪液)或低细胞和组织质量的样品(如组织针活检)。与分析型液相色谱法(色谱柱内径 2.1-5.0 毫米)相比,应用微流、毛细管或纳米液相色谱法(色谱柱内径≤ 1.0 毫米)理论上应具有更高的检测灵敏度。迄今为止,对可用于研究低体积或低质量样品的微流超高压液相色谱-质谱检测方法的研究还很有限:本文介绍了三种互补的超高效液相色谱-质谱检测方法(水样 C18 反相、脂质组学 C18 反相和亲水作用液相色谱 (HILIC)),这些方法均采用内径为 1.0 毫米的色谱柱,用于非靶向代谢组学研究。人血浆和尿液样本用于方法开发,猪血浆、尿液和泪液样本用于方法评估。收集并比较了具有相同长度、固定相和固定相粒度但两种不同柱内径(2.1 毫米和 1.0 毫米)的色谱柱的数据:结果和结论:采用 1.0 毫米内径检测时,所有三种检测方法的峰面积和峰宽都有所增加。HILIC 检测法在较低样品稀释度时具有优势,而反相 (RP) 检测法则没有任何优势。这一点可以从验证研究中看出,HILIC 分析法检测到的化合物数量要多得多。RP 检测法仍然适用于检测数百种化合物的小容量样品。总之,1.0 mm 直径色谱柱检测法适用于样品制备过程中需要稀释的小容量样品。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Development of microflow ultra high performance liquid chromatography-mass spectrometry metabolomic assays for analysis of mammalian biofluids.

Introduction and objectives: The application of untargeted metabolomics assays using ultra high performance liquid chromatography-mass spectrometry (UHPLC-MS) to study metabolism in biological systems including humans is rapidly increasing. In some of these studies there is a requirement to collect and analyse low sample volumes of biofluids (e.g. tear fluid) or low cell and tissue mass samples (e.g. tissue needle biopsies). The application of microflow, capillary or nano liquid chromatography (≤ 1.0 mm column internal diameter (i.d.)) theoretically should accomplish a higher assay sensitivity compared to analytical liquid chromatography (2.1-5.0 mm column internal diameter). To date, there has been limited research into microflow UHPLC-MS assays that can be applied to study samples of low volume or mass.

Methods: This paper presents three complementary UHPLC-MS assays (aqueous C18 reversed-phase, lipidomics C18 reversed-phase and Hydrophilic Interaction Liquid Chromatography (HILIC)) applying 1.0 mm internal diameter columns for untargeted metabolomics. Human plasma and urine samples were applied for the method development, with porcine plasma, urine and tear fluid used for method assessment. Data were collected and compared for columns of the same length, stationary phase and stationary phase particle size but with two different column internal diameters (2.1 mm and 1.0 mm).

Results and conclusions: All three assays showed an increase in peak areas and peak widths when applying the 1.0 mm i.d. assays. HILIC assays provide an advantage at lower sample dilutions whereas for reversed phase (RP) assays there was no benefit added. This can be seen in the validation study where a much higher number of compounds were detected in the HILIC assay. RP assays were still appropriate for small volume samples with hundreds of compounds being detected. In summary, the 1.0 mm i.d. column assays are applicable for small volume samples where dilution is required during sample preparation.

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来源期刊
Metabolomics
Metabolomics 医学-内分泌学与代谢
CiteScore
6.60
自引率
2.80%
发文量
84
审稿时长
2 months
期刊介绍: Metabolomics publishes current research regarding the development of technology platforms for metabolomics. This includes, but is not limited to: metabolomic applications within man, including pre-clinical and clinical pharmacometabolomics for precision medicine metabolic profiling and fingerprinting metabolite target analysis metabolomic applications within animals, plants and microbes transcriptomics and proteomics in systems biology Metabolomics is an indispensable platform for researchers using new post-genomics approaches, to discover networks and interactions between metabolites, pharmaceuticals, SNPs, proteins and more. Its articles go beyond the genome and metabolome, by including original clinical study material together with big data from new emerging technologies.
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