Electron Paramagnetic Resonance (EPR) is a sensitive and direct technique to detect, quantitate and image free radical species. In addition, the spectral modification of paramagnetic probes by tissue oxygen provides a sensitive method for in vivo oxymetry. In vivo EPR studies on intact animals is not feasible using conventional spectrometers which operate at relatively higher frequencies, where the incident radiation does not adequately penetrate the object under investigation. Because ionizing radiation ultimately results in formation of free radical species, the Radiation Biology Branch has been developing a pulsed Fourier Transform (FT) EPR system at low frequencies for spectroscopy and imaging of larger biologic objects such as tissue, tumors and even small animals. In the current configuration, samples of up to 15 cc volume can be easily accommodated for free radical detection and imaging. Feasibility of in vivo oxymetry of murine models has been demonstrated for the first time. A free radical particulate probe whose EPR spectral parameters are sensitive to tissue oxygenation has been implanted into the thigh of a mouse, and the spectra recorded repeatedly after subjecting this muscle to ischemia and reoxygenation. The spectral line width of the free radical monitored by the spectrometer validly reflected the changes in tissue oxygenation. When a tumor was grown around the site of the particulate EPR probe and EPR oxymetry performed, tumor oxygenation was found to correspond to that observed in severely ischemic tissue. In a preliminary two-dimensional imaging experiment using a murine model infused with a soluble EPR probe restricted predominantly to vasculature provided anatomical images with resolution < 0.5 mm. Futhermore the image intensity was found to be inversely proportional to breathing oxygen content. These experiments represent the first application of Fourier techniques in imaging free radicals and non-invasively deriving tissue oxygenation.
