Robust Rapid Cellular Metabolite Sensing Using Benchtop NMR and SABRE-Hyperpolarized [1-¹³C]Pyruvate.

Area of Science:

• Nuclear Magnetic Resonance (NMR) Spectroscopy
• Metabolic Engineering
• Biophysical Chemistry

Background:

• Hyperpolarized NMR significantly enhances signal sensitivity for biological dynamics.
• Signal Amplification By Reversible Exchange (SABRE) is a hyperpolarization technique.
• Investigating real-time cellular metabolism requires sensitive and rapid analytical methods.

Purpose of the Study:

• To demonstrate benchtop SABRE for preparing hyperpolarized [1-¹³C]pyruvate for yeast metabolism studies.
• To characterize real-time metabolic activity in *Saccharomyces cerevisiae* using this probe.
• To model metabolite kinetics and estimate intracellular pH changes.

Main Methods:

• Utilized Signal Amplification By Reversible Exchange (SABRE) for hyperpolarized [1-¹³C]pyruvate preparation.
• Developed a modular, compact workflow for benchtop, live-cell experiments.
• Employed high temporal resolution NMR to monitor metabolic conversions and pH dynamics.

Main Results:

• Achieved highly reproducible hyperpolarized [1-¹³C]pyruvate doses suitable for benchtop use.
• Resolved the conversion of the probe to CO₂ and bicarbonate in yeast cells over 300 seconds.
• Modeled metabolic kinetics and derived real-time pH estimates from the carbonic acid buffer system.

Conclusions:

• Benchtop SABRE-hyperpolarized [1-¹³C]pyruvate is a scalable and specific probe for metabolic phenotyping.
• The compact instrumentation is well-suited for high-throughput applications in microbial engineering and drug screening.
• This method provides dynamic metabolic screening capabilities for living cells.