Trace Phosphate Improves ZIC-pHILIC Peak Shape, Sensitivity, and Coverage for Untargeted Metabolomics

Journal of Proteome Research
Jonathan L SpaldingGary J Patti

Abstract

Existing hydrophilic interaction liquid chromatography (HILIC) methods, considered individually, each exhibit poor chromatographic performance for a substantial fraction of polar metabolites. In addition to limiting metabolome coverage, such deficiencies also complicate automated data processing. Here we show that some of these analytical challenges can be addressed for the ZIC-pHILIC, a zwitterionic stationary phase commonly used in metabolomics, with the addition of trace levels of phosphate. Specifically, micromolar phosphate extended metabolome coverage by hundreds of credentialed features, improved peak shapes, and reduced peak-detection errors during informatic processing. Although the addition of high levels of phosphate (millimolar) as a HILIC mobile phase buffer has been explored previously, such concentrations interfere with mass spectrometric (MS) detection. We show that using phosphate as a trace additive at micromolar concentrations improves analysis by electrospray MS, increasing signal for a diverse set of polar standards. Given the small amount of phosphate needed, comparable chromatographic improvements were also achieved by direct addition of phosphate to the sample during reconstitution. Our results suggest t...Continue Reading

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Citations

Dec 1, 2019·Metabolites·Luca NarduzziGaud Dervilly-Pinel
Dec 19, 2018·Cancer & Metabolism·Xiangfeng NiuGary J Patti
Feb 3, 2021·Journal of Proteome Research·Prasad PhapaleTheodore Alexandrov
Apr 11, 2020·Journal of the American Chemical Society·Miriam Sindelar, Gary J Patti

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