Rapid determination of ascorbate by ESR spectroscopy.

Optimal conditions for the determination of ascorbate, based on the detection of its radical by ESR spectroscopy, were established. The optimal pH for the measurement of the ascorbate radical was 10, whereas that for the gallate and caffeate radical was much higher. This was due to the liability of the ascorbate radical at higher pH. When the radical intensity of ascorbate vs. concentration was plotted double logarithmically, a linear correlation was established between these two parameters within the ascorbate concentration of 0.03-10 mM. ESR analysis of the homogenates under alkaline condition demonstrated that ascorbate is distributed in various mouse organs and that the ascorbate radical is unstable at neutral pH in the brain and liver, suggesting the presence of metabolizing substances. The present study suggests the applicability of the present technique for the study of ascorbate metabolism and identification of physiological factors which regulate the radical intensity of ascorbate.