Human Metabolomics, North-West University, Potchefstroom Campus, Private Bag X6001, Box 269, Potchefstroom, 2520, NW, South Africa.
Sci Rep. 2023 Oct 16;13(1):17591. doi: 10.1038/s41598-023-44690-7.
Urine is ideal for non-targeted metabolomics, providing valuable insights into normal and pathological cellular processes. Optimal extraction is critical since non-targeted metabolomics aims to analyse various compound classes. Here, we optimised a low-volume urine preparation procedure for non-targeted GC-MS. Five extraction methods (four organic acid [OA] extraction variations and a "direct analysis" [DA] approach) were assessed based on repeatability, metabolome coverage, and metabolite recovery. The DA method exhibited superior repeatability, and achieved the highest metabolome coverage, detecting 91 unique metabolites from multiple compound classes comparatively. Conversely, OA methods may not be suitable for all non-targeted metabolomics applications due to their bias toward a specific compound class. In accordance, the OA methods demonstrated limitations, with lower compound recovery and a higher percentage of undetected compounds. The DA method was further improved by incorporating an additional drying step between two-step derivatization but did not benefit from urease sample pre-treatment. Overall, this study establishes an improved low-volume urine preparation approach for future non-targeted urine metabolomics applications using GC-MS. Our findings contribute to advancing the field of metabolomics and enable efficient, comprehensive analysis of urinary metabolites, which could facilitate more accurate disease diagnosis or biomarker discovery.
尿液是进行非靶向代谢组学分析的理想样本，可深入了解正常和病理细胞过程。由于非靶向代谢组学旨在分析各种化合物类别，因此最佳的提取方法至关重要。在这里，我们优化了一种用于非靶向 GC-MS 的低容量尿液制备方法。根据重复性、代谢组覆盖范围和代谢物回收率，评估了五种提取方法（四种有机酸[OA]提取方法的变化和一种“直接分析”[DA]方法）。DA 方法具有出色的重复性，实现了最高的代谢组覆盖范围，可从多个化合物类别中检测到 91 种独特的代谢物。相比之下，OA 方法可能并不适用于所有非靶向代谢组学应用，因为它们偏向于特定的化合物类别。因此，OA 方法的局限性在于化合物回收率较低，且有更多的未检出化合物。通过在两步衍生化之间增加一个干燥步骤，DA 方法得到了进一步的改进，但并未受益于尿酶样品预处理。总体而言，本研究建立了一种使用 GC-MS 进行未来非靶向尿液代谢组学应用的改进的低容量尿液制备方法。我们的研究结果有助于推动代谢组学领域的发展，并实现对尿液代谢物的高效、全面分析，这可能有助于更准确的疾病诊断或生物标志物发现。
