Plasma L-5-oxoproline carbon and nitrogen kinetics in healthy young adults

Abstract

L-5-oxoproline (OP), an intermediate of the γ-glutamyl cycle of glutathione synthesis and degradation, may serve as a probe for the state of glutathione kinetics. We explored the whole-body carbon and nitrogen kinetics of OP in five male healthy subjects (75.2 kg; 181 cm; 26 y) after a 5-d adaptation to an adequate L-amino acid-based diet (160 mg N · kg-1 · d-1; 188 kJ · kg-1 · d-1), using a crossover design. On day 6 of the diet period, we carried out an 8-h tracer protocol (3 h fast; 5 h fed; 2/3 of dally nitrogen intake) with intravenous infusion of L-[1-13C]oxoproline and L-[3,3-2H]cysteine or, in randomized order, on the second occasion, L- [15N]oxoproline and L-[3,3-2H]cysteine. Plasma OP was isolated by cation exchange and after addition of internal standards (DL-[2H3]-5-oxoproline; L-[15N, U-13C5]-5-oxoproline; DL-[2H3]-glutamic acid) derivatized to form TBDMS esters and measured by gas chromatography/mass spectrometry. Plasma OP concentration did not differ between fed and fasted state (fast: 59.4 ± 8.3; fed 59.2 ± 8.9 nmol/mL). 13C- and 15N OP flux during the fasted and fed state were 19 ± 3.6, 21.2 ± 3.2, and 22.6 ± 3.9, 25.8 ± 4.3 μmol · kg-1 · 30 min-1, respectively. OP oxidation was 15.6 ± 3.6 and 17.9 ± 3.5 μmol · kg-1 · 30 min-1, in fasting and feeding, respectively, (P < 0.05). More than 80% of the plasma flux was oxidized. These findings are compared with the published literature on GSH turnover in plasma of human subjects and underscore the need to define more completely the dynamic aspects of glutathione metabolism and of the intermediates of the γ-glutamyl cycle.