Hydrophilic Interaction Liquid Chromatography (HILIC) chromatography is widely applied in metabolomics as a complementary strategy to reverse phase chromatography. Nevertheless, it still faces several issues in terms of peak shape and compounds ionization, limiting the automatic de-convolution and data semi-quantification performed through dedicated software. A way to improve the chromatographic and ionization performance of a HILIC method is to modify the electrostatic interactions of the analytes with both mobile and stationary phases. In this study, using a ZIC-HILIC chromatographic phase, we evaluated the performance of ammonium fluoride (AF) as additive salt, comparing its performance to ammonium acetate (AA). Three comparative criteria were selected: (1) identification and peak quality of 34 standards following a metabolomics-specific evaluation approach, (2) an intraday repeatability test with real samples and (3) performing two real metabolomics fingerprints with the AF method to evaluate its inter-day repeatability. The AF method showed not only higher ionization efficiency and signal-to-noise ratio but also better repeatability and robustness than the AA approach. A tips and tricks section is then added, aiming at improving method replicability for further users. In conclusion, ammonium fluoride as additive salt presents several advantages and might be considered as a step forward in the application of robust HILIC methods in metabolomics.

中文翻译：

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氟化铵作为基于 HILIC 的 LC-HRMS 代谢组学的合适添加剂。

亲水相互作用液相色谱（HILIC）色谱作为反相色谱的补充策略广泛应用于代谢组学中。尽管如此，它仍然面临峰形和化合物电离方面的几个问题，限制了通过专用软件执行的自动反卷积和数据半定量。提高 HILIC 方法的色谱和电离性能的一种方法是改变分析物与流动相和固定相的静电相互作用。在本研究中，我们使用 ZIC-HILIC 色谱相评估了氟化铵 (AF) 作为添加剂盐的性能，并将其性能与醋酸铵 (AA) 进行了比较。选择了三个比较标准：（1）按照代谢组学特定评估方法对 34 个标准品进行鉴定和峰质量，（2）使用真实样品进行日内重复性测试，以及（3）使用 AF 方法进行两个真实的代谢组学指纹来评估其日间重复性。与 AA 方法相比，AF 方法不仅表现出更高的电离效率和信噪比，而且具有更好的重复性和鲁棒性。然后添加提示和技巧部分，旨在为更多用户提高方法的可复制性。总之，氟化铵作为添加剂盐具有多种优势，可能被视为代谢组学中稳健的 HILIC 方法应用的一个进步。