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Measuring glutathione fractional synthesis in vivo with deuterated water
Author(s) -
Cabral Carolina B,
Bullock Kevin,
Bischoff David,
Yu YongMing,
Tompkins Ronald G,
Kelleher Joanne K
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a1470-a
Subject(s) - glutathione , chemistry , kinetics , deuterium , amino acid , in vivo , tracer , cysteine , intraperitoneal injection , isotope , radiochemistry , chromatography , biochemistry , endocrinology , biology , physics , microbiology and biotechnology , nuclear physics , enzyme , quantum mechanics
Previous stable isotope methods for estimating the synthesis of glutathione have utilized a labeled version of one of the three amino acids with the assumption that the precursor enrichment of the amino acid in plasma is equivalent to the precursor enrichment at the site of synthesis. We developed a method for estimating GSH synthesis using deuterated water (D 2 O). New Zealand rabbits received intraperitoneal injection of deuterated saline calculated to result in 4–5% enrichment of total body water. Drinking water was enriched to 5% D 2 O. Blood samples were drawn and the experiment was terminated at various times over a two‐week period. Liver samples were obtained at the termination of the experiment. Hepatic and erythrocyte GSH labeling kinetics were obtained by LC/MS using the N ethylmaleimide derivative of GSH. When body water was 4.7% D 2 O, the labeling of erythrocyte and hepatic GSH increased to a plateau value corresponding to a 14% enrichment at M+1. This corresponds to the exchange of 6–7 of the 10 exchangeable H with the D enriched water during synthesis. We developed a probability‐based model to estimate the exchangeable H. Labeling kinetics indicated that the fractional synthesis rate of erythrocyte GSH was 4.3 %/ day, identical to the value found with 13 C cysteine as tracer. Our studies indicate that GSH synthesis from D 2 O could be estimated in human subjects at D 2 O enrichment of 1%. The advantages of D 2 O to estimate glutathione synthesis are that no assumptions of amino acid labeling are required and the kinetics maybe evaluated over many days noninvasively and inexpensively. Supported by NIH P50GM021700, RO1ES013925 and Shriners 8740.