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Trace Phosphate Improves ZIC-pHILIC Peak Shape, Sensitivity, and Coverage for Untargeted Metabolomics
Jonathan L Spalding1,2,3, Fuad J Naser1, Nathaniel G Mahieu1
1Department of Chemistry , Washington University in St. Louis , St. Louis , MO 63130 , United States.
Journal of Proteome Research
|August 31, 2018
Summary
Adding trace phosphate to hydrophilic interaction liquid chromatography (HILIC) significantly enhances polar metabolite analysis in untargeted metabolomics, improving data quality and coverage.
Area of Science:
- Analytical Chemistry
- Biochemistry
- Metabolomics
Background:
- Hydrophilic interaction liquid chromatography (HILIC) methods individually show poor performance for many polar metabolites, limiting metabolome coverage and complicating data processing.
- Existing HILIC methods struggle with a significant fraction of polar metabolites, hindering comprehensive analysis in fields like metabolomics.
Purpose of the Study:
- To improve the chromatographic performance of ZIC-pHILIC, a common HILIC stationary phase used in metabolomics.
- To enhance metabolome coverage and reduce data processing errors by optimizing HILIC methods for polar metabolites.
Main Methods:
- Investigated the effect of trace phosphate addition (micromolar concentrations) to the mobile phase of ZIC-pHILIC chromatography.
- Evaluated the impact of phosphate on metabolome coverage, peak shape, peak detection, and mass spectrometric (MS) signal intensity.
- Compared direct sample reconstitution with phosphate versus mobile phase addition.
Main Results:
- Micromolar phosphate addition extended metabolome coverage by hundreds of features and improved peak shapes.
- Phosphate addition reduced peak-detection errors during automated data processing.
- Trace phosphate enhanced electrospray mass spectrometry (MS) signal for polar standards without interference.
Conclusions:
- Defects in ZIC-pHILIC performance are primarily due to electrostatic interactions, which can be modulated by trace phosphate.
- Micromolar phosphate addition offers a methodological improvement for untargeted metabolomics, enhancing coverage and data quality.
- Understanding these electrostatic interactions advances knowledge of HILIC limitations and analytical strategies.
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