A study was made of the in vivo detectability of a pH-sensitive, imidazolidine spin probe, and the efficacy of low-frequency electron spin resonance (ESR)-based techniques for pH measurement in vitro and in vivo in rats. The techniques used were longitudinally-detected ESR (LODESR) and field-cycled dynamic nuclear polarization (FC-DNP) for in vitro and in vivo measurements, and radiofrequency (RF)-and X-band ESR for comparisons in vitro. The spin probe was hexamethyl imidazolidine (HMI) with a pK of 4.6. All techniques detected HMI. Detection by FC-DNP implies coupling between the free radical and solvent water spins. Separations between the three spectral lines of the nitroxide radical, relative to measurement frequency, were consistent with theory. The overall spectrum width from unprotonated HMI (pH > pK) was greater than that from protonated agent (pH < pK). This was observed in vitro and in vivo. Longer-term studies showed that HMI is detectable and has the same spectral width (i.e., is at the same pH) up to 2 hr after gavage into the stomach, although the magnitude of the signal decreases rapidly during the first hour. The imidazoline and imidazolidine nitroxide radicals show a reversible pH effect in their electron spin resonance (ESR) spectra. This was described in 1982 (1,2), when it was demonstrated that the N-3 atom of the ring can take up a proton in an acidic environment, resulting in a change in the unpaired electron density at the N-1 atom (3) (see Fig. 1). Studies of this effect were recently reviewed by Khramtsov and Volodarsky (4). The compound 2,2,3,4,5,5-hexamethylimidazolidine-1-yloxy (referred to here as HMI), when studied by ESR spectroscopy at Xband (9.2 GHz) or lower frequencies, has a smaller hyperfine coupling constant in an acid environment (i.e., when protonated) than it does in a more alkaline environment. When measured by ESR, this effect demonstrates a pK (the pH at which half the agent is protonated) of about 4.6 (5). Khramtsov and Volodarsky (4) showed that at high measurement frequencies (140 GHz), at pH 4.7, the spectrum from HMI consists of two partially superimposed triplets: one deriving from the protonated HMI(H ϩ ) state, and one from the unprotonated state. At lower frequencies (X-band and below), however, the triplets overlap more closely and appear as single low-and mid-field lines with a partial doubling of the high-field line in the X-band spectrum (4). At 1.1 GHz there is a broadening of both high-and lowfield lines (6). When the spectrum is observed at a pH value other than its pK, one of the triplets dominates and the spectrum appears as a simple triplet with a narrower splitting for HMI in acid solution and a wider splitting at higher pH. The relative proportion of the two triplets making up the spectrum varies with pH over a substantial range, permitting the effect to be used as an ESR-based pH meter (1). The effect has been observed to occur at very low measurement frequencies (280 MHz) in solutions of HMI studied in vitro (5), and may therefore be of value for...

No abstract

A novel imaging modality is introduced which uses radiofrequency longitudinally detected electron spin resonance (RF-LODESR). It is capable of providing qualitative and semiquantitative information on a variety of parameters reflecting physiological function, the most significant being tissue oxygenation. Effective spin-lattice (T * 1e ) and spin-spin (T * 2e ) electronic relaxation time maps of the abdomen of living 200-g rats were generated after intravenous administration of a triarylmethyl free radical (TAM). These maps were used to evaluate oxygen distribution. Differences between the liver, kidneys, and bladder were noted. Conclusions were made regarding the distribution, perfusion, and excretion rate of the contrast medium. Ligature-induced anoxia in the kidney was also visualized. LODESR involves transverse ESR irradiation with a modulated excitation, and observing oscillations in the spin magnetization parallel to the main magnetic field. The T * 1e and T * 2e maps were calculated from a set of LODESR signal phase images collected at different detection frequencies. Each phase image also provides qualitative information on tissue oxygen levels without any further processing. This method presents an alternative to the conventional transverse ESR linewidth-based oximetry methods, particularly for animal whole-body imaging applications. Magn