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Rats were given vitamin E (Vit-E), idebenone (ID), or vitamin C (Vit-C) in their food for 2 or 4 weeks. After feeding, the ability of rats to reduce 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (Tempol) in terms of the half-life of Tempol was examined as a specific marker. Tempol was repeatedly injected intravenously, and its half-life was serially evaluated by an in vivo electron spin resonance (ESR) technique. The radical-reducing ability in rats was enhanced differently by Vit-E, ID, and Vit-C, i.e., slow onset of the ability after Vit-E and ID (lipid-soluble antioxidants) and fast onset after Vit-C (a water-soluble antioxidant).

The flexible surface-coil-type resonator (FSCR) operating in a 700 MHz microwave electron spin resonance (ESR) system was applied to measure the nitroxide radicals at a specific area in rats. The FSCR was composed of a single-turn coil with a diameter of 5 mm and two flexible coaxial lines of 450 mm in length. For an endoscope-like application, the FSCR was inserted into the rectum of the rat and sequential changes in the ESR signals of the intravenously injected nitroxide radical (4-hydroxy-2,2,6,6-tetramethyl-piperidine-1-oxyl; TEMPOL or 3-carbamoyl-2,2,5,5-tetramethylpyrrolidine-1-yloxy; carbamoyl-PROXYL) were measured. The ESR signal intensity of the nitroxide decreased according to first-order kinetics. For a stethoscope-like application, the FSCR was placed at several sites on the abdominal skin of the rats receiving a subcutaneous injection of carbamoyl-PROXYL, and diffusion and/or metabolism of the radical in the skin was observed.

A rapid field scan L-band electron spin resonance computed tomography (ESR-CT) system has been developed in order to obtain three-dimensional images of the distribution of free radicals in a sample. The ESR system comprises an L-band (ca. 700 MHz) ESR spectrometer, an air-core electromagnet equipped with a field-scanning coil, three field-gradient coils, and a microcomputer.The rapid field scan (15 mT/s) causes a delay in the magnetic-field strength due to eddy-current losses. However, it was found that the magnetic field strength could be corrected by a simple mathematical equation related to a low-pass filter model. Using this system, the original ESR data used to obtain one three-dimensional ESR-CT image could be obtained in 150 s. The time course of the ESR-CT images of the test sample containing the reaction of nitroxide radical with ascorbate ion has been shown as an example of a spatiotemporal measurement.

The rate constants of nitroxide reduction by ascorbate ion were determined from a time-series measurement of ESR images, that is, ESR spatiotemporal images, which were measured with a rapid field scan L-band ESR-CT system. Two test tubes containing the phosphate buffer solutions of 3-carbamoyl-2,2,5,5-tetramethylpyrrolidin-l-yloxy (C-PROXYL) and ascorbate ion with different concentrations were placed together in the loop-gap resonator and a time-series measurement of 2-dimensional ESR images were obtained. The first-order rate constants for each reduction were determined from the time course of the total gray scale at each test tube region. These rate constants agreed with those measured individually from L-band or X-band ESR spectra. In order to study the dynamics of free radicals at an interested region in a living biological sample, it is very important to obtain the time course of radical concentration at the interested region in the sample in vivo. For this purpose, fast electron spin resonance (ESR) imaging is required. We have previously developed the rapid field scan L-band ESR computed tomography (ESR-CT) system equipped with an air-core electromagnet.' This system has been applied to biological samples and the spatiotemporal ESR images of nitroxide distribution have been displayed.2,3 However, no quantitative analysis, such as the determination of rate constants for the nitroxide reduction has yet been carried out.Recently a new data processing system has been developed by Yuasa and his coworkers4 to obtain ESR images and each gray scale value of an interested region in a series of ESR images measured is studied. In this study, the rate constants for nitroxide radical reduction are determined by using the rapid field scan L-band ESR-CT system with the data processing program. First, a linear relationship is confirmed between the gray scale value of the image and the nitroxide radical concentration. After this confirmation, the first-order rate constants of nitroxide radical reduction by ascorbate ion were determined from the time course of the gray scale value. Experimental L-Band ESR-CT systemThe L-band ESR-CT system used in this study is composed of a rapid field scan L-band ESR imaging unit and a data-processing system. The details of the L-band ESR imaging unit have been described in a previous paper.' For a 2-dimensional ESR image, a field gradient was applied at 1 mT/cm changing its directions in 10-degree steps, and data from nineteen (18+1) spectra were obtained. In processing the data, the low-pass filter technique (DLPF) and the simultaneous iterative reconstruction technique (SIRT) were used to deconvolute the ESR spectral data and to reconstruct a 2-dimensional ESR image with 64X64 points (50X50 mm), respectively.4 In illustrating a picture of a series of ESR images, the maximum value of the gray scale was taken as 100%. Furthermore, a region of interest was selected from an initial image in a series of ESR images and a total gray scale value of the selected region was calculated for each s...

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