Scanning central carbon metabolism: a HILIC-HR-TOF-MS metabolome method

Area of Science:

• Metabolomics
• Analytical Chemistry
• Cellular Metabolism

Background:

• Central carbon metabolism (CCM) is vital for cellular energy and biosynthesis.
• Analytical tools are needed to quantify CCM intermediates for research.
• Understanding CCM is crucial for biomedical and biotechnological applications.

Purpose of the Study:

• To develop a liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method for CCM analysis in mammalian cells.
• To enable semi-quantification of CCM metabolites.
• To provide a robust platform for studying metabolic alterations.

Main Methods:

• One-step liquid extraction for hydrophilic metabolites.
• Stable isotope dilution with a U-¹³C-yeast internal standard (IS).
• Hydrophilic interaction liquid chromatography (HILIC) coupled to Zeno-time-of-flight (ZenoTOF) MS for LC-HRMS analysis.

Main Results:

• Successfully identified and semi-quantified 82 CCM metabolites, with 77 confirmed by standards.
• Internal standard normalization improved method robustness and metabolite semi-quantification.
• The HILIC-HR-TOF-MS method effectively captured metabolic changes induced by chemical inhibitors.

Conclusions:

• The developed HILIC-HR-TOF-MS method is efficient for mapping CCM metabolic intermediates in mammalian cells.
• This approach has broad potential for analyzing diverse biological samples.
• Applications include drug development and various biomedical research areas.