EPR imaging of free radicals in a coffee bean

Electron paramagnetic Resonance (EPR) imaging is a non-invasive and quantitative methodology with applications in materials and biomedical research.

Materials science applications include detection and imaging of endogenous free radicals, with most experiments being carried out a X-band (ca. 9.5 GHz).

In biomedical research, EPR imaging is able to determine tissue microenvironment parameters including oxygen partial pressure, pH, the redox microenvironment and reactive oxygen species (ROS) in vivo. Due to the lossy aqueous environment, imaging is carried out at L-band (ca. 1 GHz) and is applicable to small animals, e.g. mouse.

A rich variety of suitable paramagnetic probes for EPR exist: they have simple EPR spectra, can be administered at levels that are well tolerated, and have a pharmacologic half-life that is longer than the spectroscopy/imaging time.

Partial pressure of oxygen pO2: Although dissolved molecular oxygen is paramagnetic, it cannot be detected directly by EPR. The pO2-imaging capability in EPR stems from the fact that the paramagnetic molecular oxygen affects the relaxation rates of an administered paramagnetic probe, thus causing spectral broadening that is proportional to pO2.

Measurement of pH: This measurement is based on the incorporation of ionisable groups into the structure of the triarylmethyl radical. pH measurement from 5.6 to 8.2 is possible by determining the ratio of the EPR spectrum from protonated and deprotonated TAM molecules.

Spin-trapping techniques: Reactive oxygen species, such as OH•, O2¯•, ROO•, RO•, CH3O• and CH3• play a critical role in many biological processes. Direct detection of these species by EPR is typical not possible as the radicals are short lived and do not accumulate. To circumvent this, the ROS are ‘trapped’ or stabilised by addition of a radical spin trap (e.g. DMPO) which forms a stable adduct with e.g. a OH• radical, which persists long enough for the spin trap adduct to be measured by EPR. This allows the concentration of the ROS to be followed over time, and by examination of the spin trap adduct spectrum the type of ROS can be identified in suitable cases.