Glutathione concentration was assessed using a Glutathione Assay Kit (Sigma-Aldrich). Cultured cells and mouse kidneys were harvested and lysed by repeated freeze–thaw cycles in 5% 5-sulfosalicylic acid solution and centrifuged at 10,000 r.p.m. for 10 min. A total of 10 μl of supernatant from each sample was added to 150 μl of the working mixture (glutathione reductase and DTNB solution) and incubated for 5 min at RT. NADPH solution was added to each well, and glutathione levels were determined by kinetic absorbance measurement (1 min intervals for 5 min) at a wavelength of 412 nm. The glutathione content of each specimen was calculated by comparison with the standard.
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