Reduced glutathione (GSH) estimation

A homogenate of ten-day-old seedlings (0.5 g) was prepared in 2.5 ml of 5% (v/v) TCA. Homogenate was centrifuged at 4,000xg for 10 min. The supernatant (0.1 ml) was used for the estimation of GSH. The supernatant (0.1 ml) was made up to 1.0 ml using 0.2 M sodium phosphate buffer (pH 8.0) followed by the addition of 2.0 ml of freshly prepared DTNB (5,5′- dithiobis (2-nitrobenzoic acid) solution. The concentration of GSH was determined spectrophotometrically at 412 nm after 10 min of incubation (Moron et al., 1979). The values are expressed as µmoles g ⁻¹ sample.