In vivo monitoring of hepatic glutathione in anesthetized rats by 13C NMR

A method for in vivo 13C NMR monitoring of hepatic glutathione (GSH) in intact, anesthetized rats has been developed. Studies were conducted using a triple‐tuned, surgically implanted surface coil designed for this animal model. The coil permitted complete decoupling and sufficient resolution in the 13C NMR spectrum to monitor the time course of hepatic 13C‐metabolites of intravenously administered 2‐13C‐glycine, particularly GSH at 44.2 ppm and serine signals at 61.1 and 57.2 ppm, respectively. It further allowed concomitant monitoring of high‐energy phosphagens and intracellular pH by 31P NMR. To confirm in vivo NMR peak assignments, we compared high‐resolution 2D 1H{13C} heteronuclear multiple quantum coherence and 1D 13C spectra of hepatic perchloric acid extracts to those of authentic standards. The fractional isotopic enrichment of hepatic 13C‐glycine increased exponentially at a rate of 1.68 h−1 and reached its plateau level of 81% in 2 h. The 13C fractional isotopic enrichment of GSH increased exponentially at a rate of 0.316 h−1 and reached 55% after 4 h of 2‐13C‐glycine infusion, but without achieving a plateau. To confirm that the resonance at 44.2 ppm resulted from GSH, a rat was given an intravenous dose of 2‐oxothiazolidine‐4‐carboxylic acid (OTC), a cysteine precursor that increases intracellular GSH. As expected, with OTC administration the hepatic 13C GSH‐to‐glycine peak area increased more than sevenfold. Magn Reson Med 48:430–439, 2002. © 2002 Wiley‐Liss, Inc.