The NMR signal of reduced glutathione (GSH) was monitored in intact human erythrocytes by the 1H spin-echo Carr-Purcell-Meiboom-Gill pulse sequence. Addition of GSH, which was unable to cross the erythrocyte membrane, produced an approximate two-fold increase of the GSH signal in glucose-depleted cells. Addition of oxidised glutathione (GSSG), did not affect the signal, and addition of GSH to hemolysates gave a much smaller increase. Reduction of internal GSSG by NADPH-dependent enzymes was excluded by experiments with glucose-supplied or glucose-6-phosphate dehydrogenase deficient cells. Involvement of external thiol groups of the erythrocyte membrane was shown by the lack of effect in cells treated with an impermeable thiol-blocking compound. Involvement of spectrin was indicated by the proportional loss of the effect in erythrocytes with variable genetic deficiency of spectrin. Protein-glutathione mixed disulfides appeared to be the source of the NMR response since an increase of their content, by diamide treatment or aging procedures, produced a higher GSH signal, while their reduction by permeable reductants gave the opposite effect. It is concluded that GSH can transduce its reducing power by a thiol/disulfide exchange mechanism that sequentially involves sulfur-rich proteins spanning across the erythrocyte membrane.