不同粪便提取方法对人类粪便样本代谢组学测量的评价。

IF 3.3 Q2 NUTRITION & DIETETICS
BMJ Nutrition, Prevention and Health Pub Date : 2021-07-02 eCollection Date: 2021-01-01 DOI:10.1136/bmjnph-2020-000202
Vanessa Erben, Gernot Poschet, Petra Schrotz-King, Hermann Brenner
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引用次数: 0

摘要

背景:人类粪便样本的代谢组学分析在生物医学研究中有着广泛的应用,包括结肠直肠肿瘤的早期检测。然而,由于代谢物的复杂性,对于如何处理粪便代谢组学测量样品以获得广泛的亲水和疏水物质尚无共识。方法:我们使用健康研究参与者的冷冻粪便样本(50毫克)。解冻后的粪便样本使用八种不同的处理方案和不同的溶剂(溶剂如磷酸盐缓冲盐水、异丙醇、甲醇、乙醇、乙腈和溶剂混合物,有或没有蒸发和浓缩步骤)进行处理。随后使用MxP Quant 500试剂盒(Biocrates)测量代谢物。最佳方案随后被用于比较具有不同饮食习惯的参与者的粪便样本。结果:在这项研究中,我们能够确定多达340种不同化学类别的代谢物,这些代谢物是从8种不同方案的健康研究参与者的粪便样本中提取的。极性代谢物,如氨基酸,可以用每种方法测量,而其他代谢物类别,特别是脂类(在干燥步骤后使用异丙醇和乙醇或甲醇更好),更依赖于所用溶剂或溶剂组合。在人类粪便中只检测到少量的甘油三酯或酰基肉碱。使用异丙醇(131个代谢物>检出限(LOD))或使用乙醇或甲醇和甲基叔丁基醚(MTBE)包括蒸发和浓缩步骤(分别为303和342个代谢物>检出限)的方案的提取效率高于其他方案。我们在素食者、半素食者和非素食者之间发现了显著的粪便代谢物差异。结论:对于粪便样本中代谢物的评估,我们发现使用溶剂如异丙醇和使用乙醇或甲醇的方案,以及包括蒸发和浓缩步骤的MTBE在检测不同化学类别代谢物的数量方面优于本研究中测试的其他方案。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Evaluation of different stool extraction methods for metabolomics measurements in human faecal samples.

Evaluation of different stool extraction methods for metabolomics measurements in human faecal samples.

Evaluation of different stool extraction methods for metabolomics measurements in human faecal samples.

Background: Metabolomics analysis of human stool samples is of great interest for a broad range of applications in biomedical research including early detection of colorectal neoplasms. However, due to the complexity of metabolites there is no consensus on how to process samples for stool metabolomics measurements to obtain a broad coverage of hydrophilic and hydrophobic substances.

Methods: We used frozen stool samples (50 mg) from healthy study participants. Stool samples were processed after thawing using eight different processing protocols and different solvents (solvents such as phosphate-buffered saline, isopropanol, methanol, ethanol, acetonitrile and solvent mixtures with or without following evaporation and concentration steps). Metabolites were measured afterwards using the MxP Quant 500 kit (Biocrates). The best performing protocol was subsequently applied to compare stool samples of participants with different dietary habits.

Results: In this study, we were able to determine up to 340 metabolites of various chemical classes extracted from stool samples of healthy study participants with eight different protocols. Polar metabolites such as amino acids could be measured with each method while other metabolite classes, particular lipid species (better with isopropanol and ethanol or methanol following a drying step), are more dependent on the solvent or combination of solvents used. Only a small number of triglycerides or acylcarnitines were detected in human faeces. Extraction efficiency was higher for protocols using isopropanol (131 metabolites>limit of detection (LOD)) or those using ethanol or methanol and methyl tert-butyl ether (MTBE) including an evaporation and concentration step (303 and 342 metabolites>LOD, respectively) than for other protocols. We detected significant faecal metabolite differences between vegetarians, semivegetarians and non-vegetarians.

Conclusion: For the evaluation of metabolites in faecal samples, we found protocols using solvents like isopropanol and those using ethanol or methanol, and MTBE including an evaporation and concentration step to be superior regarding the number of detected metabolites of different chemical classes over others tested in this study.

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来源期刊
BMJ Nutrition, Prevention and Health
BMJ Nutrition, Prevention and Health Nursing-Nutrition and Dietetics
CiteScore
5.80
自引率
0.00%
发文量
34
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