Despite reports of lower glutathione (GSH) concentration in sickle cell disease (SCD), the in vivo kinetic mechanism(s) responsible for GSH deficiency is unknown. To determine whether suppressed synthesis was responsible for the lower erythrocyte GSH concentration, we used a prime intermittent infusion of ²H₂glycine to measure erythrocyte GSH synthesis in vivo in 23 individuals with homozygous ^s sickle cell disease and in 8 healthy controls. Erythrocyte cysteine concentration, the rate limiting precursor for GSH synthesis, plasma markers of oxidant damage and dietary intakes of energy and protein were also measured Compared to values of controls, subjects with SCD had significantly lower erythrocyte GSH (p<0.04) and cysteine concentrations (p<0.004) but significantly faster fractional rates of GSH synthesis (p<0.02). The absolute rates of GSH synthesis in subjects with SCD compared with control subjects was greater by ~57% (p=0.062). However, the concentrations of markers of oxidative damage, plasma derivatives of reactive oxygen metabolites, plasma nitrotyrosine, urinary isoprostane:creatinine ratio, and GSH:GSSG ratio as well as dietary intakes of energy, protein and GSH precursor amino acids, were not different between subjects with SCD and controls. The findings of this study suggest that the lower erythrocyte GSH of SCD patients is not due to suppressed synthesis or impaired regeneration, but rather to increased consumption. In addition, the lower erythrocyte cysteine concentration plus the faster rate of GSH synthesis strongly suggest that the endogenous cysteine supply is not sufficient to meet all anabolic demands; hence cysteine may be a conditionally essential amino acid in individuals with SCD.