Intracellular pH as measured by 19F NMR

Area of Science:

• Biophysical Chemistry
• Nuclear Magnetic Resonance Spectroscopy
• Cellular Biology

Background:

• Phosphorus-31 Nuclear Magnetic Resonance (31P NMR) is a standard method for intracellular pH (pHi) measurement.
• However, 31P NMR is not suitable for all biological systems, limiting pHi analysis in certain contexts.
• Developing alternative, non-destructive methods for pHi measurement is crucial for comprehensive biological research.

Purpose of the Study:

• To introduce and validate the use of fluorine-19 Nuclear Magnetic Resonance (19F NMR) with novel fluorinated pH indicators for pHi measurements.
• To demonstrate the advantages of 19F NMR over 31P NMR in specific applications.
• To explore the potential for targeted, in situ generation of pH indicators within specific cellular or tissue compartments.

Main Methods:

• Utilized a series of fluorinated pH indicators for 19F NMR spectroscopy.
• Performed pH measurements in various biological systems where 31P NMR is impractical.
• Investigated the use of indicator precursor molecules designed for in situ generation via cellular enzymes.

Main Results:

• 19F NMR pH measurements were found to be rapid, sensitive, and compatible with commercial NMR instruments.
• The 19F NMR method successfully extended non-destructive pH measurements to systems unsuitable for 31P NMR.
• The use of exogenous indicators was shown to be advantageous for independent confirmation of pHi values and tracking of weak acid distribution.
• Demonstrated the feasibility of designing indicator precursors for enzyme-mediated in situ generation.

Conclusions:

• Fluorinated pH indicators and 19F NMR provide a powerful, versatile tool for non-destructive pHi determination.
• This approach complements and expands upon existing methods like 31P NMR.
• Future applications include precise, compartment-specific pHi measurements through targeted in situ indicator generation.