BACKGROUND High-throughput simultaneous quantitative and qualitative (Quan/Qual) analysis is attractive to combine targeted with non-targeted analysis, e.g. in pharmacometabolomics and drug metabolism studies. This study aimed to investigate the possibilities and limitations of high-throughput Quan/Qual analysis by ultra-high performance liquid chromatography (UHPLC) coupled with high-resolution mass spectrometry (HRMS), to develop a widely applicable Quan/Qual UHPLC-HRMS method and to provide recommendations for Quan/Qual method development. METHODS A widely applicable 4.25-min UHPLC method for small-molecules was used to investigate and optimize mass spectrometric parameters of a Synapt G2S for Quan/Qual analysis. The method was applied on a rat metabolomics study investigating the effect of the fasting state and administration of a dosing vehicle on the rat plasma metabolic profile. RESULTS Highly important parameters for high-throughput Quan/Qual analysis were the scan mode and scan rate. A negative correlation was found between the amount of qualitative information that a method can provide and its quantitative performance (accuracy, precision, sensitivity, linear dynamic range). The optimal balance was obtained using the MSE scan mode with a short scan time of 30 ms. This 4.25-min Quan/Qual analysis method enabled quantification with accuracy and precision values ≤ 20% at the lowest quality control (QC) level and ≤15% at higher QC levels for 16 out of 19 tested analytes. It provided both parent m/z values and fragmentation spectra for compound identification with limited loss of chromatographic resolution and it revealed biologically relevant metabolites in its application to the metabolomics study. CONCLUSION Quan/Qual method development requires balancing between the amount of qualitative data, the quality of the quantitative data and the analysis time. Recommendations are provided for MS resolution, scan mode, scan rate, smoothing and peak integration in Quan/Qual method development and analysis.

中文翻译：

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使用液相色谱与四极杆飞行时间质谱联用进行 Quan/Qual 分析的质谱建议

背景高通量同时定量和定性（Quan/Qual）分析对于将靶向分析与非靶向分析相结合是有吸引力的，例如在药物代谢组学和药物代谢研究中。本研究旨在探讨超高效液相色谱 (UHPLC) 与高分辨率质谱 (HRMS) 联用进行高通量 Quan/Qual 分析的可能性和局限性，以开发一种广泛适用的 Quan/Qual UHPLC-HRMS 方法并为 Quan/Qual 方法开发提供建议。方法 广泛适用的 4.25 分钟 UHPLC 小分子方法用于研究和优化用于 Quan/Qual 分析的 Synapt G2S 的质谱参数。该方法应用于大鼠代谢组学研究，研究禁食状态和给药载体给药对大鼠血浆代谢特征的影响。结果 高通量 Quan/Qual 分析的非常重要的参数是扫描模式和扫描速率。发现方法可以提供的定性信息量与其定量性能（准确度、精密度、灵敏度、线性动态范围）之间存在负相关。使用具有 30 ms 短扫描时间的 MSE 扫描模式获得最佳平衡。这种 4.25 分钟的 Quan/Qual 分析方法能够对 19 种测试分析物中的 16 种进行定量，准确度和精密度值在最低质量控制 (QC) 水平下≤ 20%，在较高质量控制水平下≤15%。它为化合物鉴定提供母体 m/z 值和碎裂谱图，色谱分辨率损失有限，并揭示了将其应用于代谢组学研究的生物学相关代谢物。结论 Quan/Qual 方法开发需要在定性数据量、定量数据质量和分析时间之间取得平衡。为 Quan/Qual 方法开发和分析中的 MS 分辨率、扫描模式、扫描速率、平滑和峰积分提供了建议。