FIGURE 5.

Reduction of GSSG, M-45, and M-90 by glutathione reductase. 20 μm GSSG, M-45, or M-90 were added to 22 nm glutathione reductase in phosphate buffer (100 mm, pH 7.5) containing 1 mm EDTA. After a 5-min incubation, the reaction was initiated by adding 500 μm NADPH (B1–B3, glutathione reductase system) or buffer (A1–A3, control). The reactions were terminated at 10 min by adding 20% trifluoroacetic acid to reach pH 2. The reaction mixtures were then filtered through a 3-kDa cutoff spin filter to remove macromolecules. The resulting reaction mixtures were analyzed by LC/MS.