PDF(Pubmed)
Abstract：
BACKGROUND: Accurately identifying and quantifying polar metabolites using untargeted metabolomics has proven challenging in comparison to mid to non-polar metabolites. Hydrophilic interaction chromatography and gas chromatography-mass spectrometry are predominantly used to target polar metabolites.
OBJECTIVE: This study aims to demonstrate a simple one-step extraction combined with liquid chromatography-mass spectrometry (LC-MS) that reliably retains polar metabolites.
METHODS: The method involves a MilliQ + 10% trichloroacetic acid extraction from 6 healthy individuals serum, combined with porous graphitic carbon liquid chromatography-mass spectrometry (LC-MS). The coefficient of variation (CV) assessed retention reliability of polar metabolites with logP as low as - 9. QreSS (Quantification, Retention, and System Suitability) internal standards determined the method\'s consistency and recovery efficiency.
RESULTS: The method demonstrated reliable retention (CV < 0.30) of polar metabolites within a logP range of - 9.1 to 5.6. QreSS internal standards confirmed consistent performance (CV < 0.16) and effective recovery (70-130%) of polar to mid-polar metabolites. Quality control dilution series demonstrated that ~ 80% of annotated metabolites could be accurately quantified (Pearson\'s correlation coefficient > 0.80) within their concentration range. Repeatability was demonstrated through clustering of repeated extractions from a single sample.
CONCLUSIONS: This LC-MS method is better suited to covering the polar segment of the metabolome than current methods, offering a reliable and efficient approach for accurate quantification of polar metabolites in untargeted metabolomics.
OBJECTIVE: This study aims to demonstrate a simple one-step extraction combined with liquid chromatography-mass spectrometry (LC-MS) that reliably retains polar metabolites.
METHODS: The method involves a MilliQ + 10% trichloroacetic acid extraction from 6 healthy individuals serum, combined with porous graphitic carbon liquid chromatography-mass spectrometry (LC-MS). The coefficient of variation (CV) assessed retention reliability of polar metabolites with logP as low as - 9. QreSS (Quantification, Retention, and System Suitability) internal standards determined the method\'s consistency and recovery efficiency.
RESULTS: The method demonstrated reliable retention (CV < 0.30) of polar metabolites within a logP range of - 9.1 to 5.6. QreSS internal standards confirmed consistent performance (CV < 0.16) and effective recovery (70-130%) of polar to mid-polar metabolites. Quality control dilution series demonstrated that ~ 80% of annotated metabolites could be accurately quantified (Pearson\'s correlation coefficient > 0.80) within their concentration range. Repeatability was demonstrated through clustering of repeated extractions from a single sample.
CONCLUSIONS: This LC-MS method is better suited to covering the polar segment of the metabolome than current methods, offering a reliable and efficient approach for accurate quantification of polar metabolites in untargeted metabolomics.
摘要：
背景：与中至非极性代谢物相比，使用非靶向代谢组学准确鉴定和定量极性代谢物已被证明具有挑战性。亲水相互作用色谱法和气相色谱-质谱法主要用于靶向极性代谢物。
目的：本研究旨在展示一种简单的一步提取结合液相色谱-质谱（LC-MS）的方法，该方法可靠地保留了极性代谢物。
方法：该方法涉及从6名健康个体血清中提取MilliQ+10%三氯乙酸，结合多孔石墨碳液相色谱-质谱(LC-MS)。变异系数(CV)评估了logP低至-9的极性代谢物的保留可靠性。QreSS(量化，保留,和系统适宜性）内部标准确定了方法的一致性和回收效率。
结果：该方法证明了极性代谢物在-9.1至5.6的logP范围内的可靠保留（CV<0.30）。QreSS内标证实了极性至中极性代谢物的一致性能(CV<0.16)和有效回收率(70-130%)。质量控制稀释系列表明，约80％的注释代谢物可以在其浓度范围内准确定量（皮尔逊相关系数>0.80）。通过从单个样品中重复提取的聚类证明了可重复性。
结论：这种LC-MS方法比目前的方法更适合覆盖代谢组的极性部分，为非靶向代谢组学中极性代谢物的准确定量提供了可靠和有效的方法。
目的：本研究旨在展示一种简单的一步提取结合液相色谱-质谱（LC-MS）的方法，该方法可靠地保留了极性代谢物。
方法：该方法涉及从6名健康个体血清中提取MilliQ+10%三氯乙酸，结合多孔石墨碳液相色谱-质谱(LC-MS)。变异系数(CV)评估了logP低至-9的极性代谢物的保留可靠性。QreSS(量化，保留,和系统适宜性）内部标准确定了方法的一致性和回收效率。
结果：该方法证明了极性代谢物在-9.1至5.6的logP范围内的可靠保留（CV<0.30）。QreSS内标证实了极性至中极性代谢物的一致性能(CV<0.16)和有效回收率(70-130%)。质量控制稀释系列表明，约80％的注释代谢物可以在其浓度范围内准确定量（皮尔逊相关系数>0.80）。通过从单个样品中重复提取的聚类证明了可重复性。
结论：这种LC-MS方法比目前的方法更适合覆盖代谢组的极性部分，为非靶向代谢组学中极性代谢物的准确定量提供了可靠和有效的方法。
